PLANT AMINO ACID BIOSYNTHETIC ENZYMES

Abstract

an isolated nucleic acid fragment encoding a plant cysteine γ synthase. The invention also relates to the construction of a chimeric gene encoding all or a portion of the plant cysteine γ synthase, in sense or antisense orientation, wherein expression of the chimeric gene results in production of altered levels of the plant biosynthetic enzyme in a transformed host cell.

Description

[0001]This application is a continuation-in-part of application Ser. No. 09/424,976 filed on Dec. 2, 1999 which is a national stage application of PCT/US98/12073 with an International filing date of Jun. 11, 1998, which in turn claims priority benefit of U.S. Provisional Application No. 60/049,406, filed Jun. 12, 1997 and U.S. Provisional Application No. 60/065,385, filed Nov. 12, 1997.

FIELD OF THE INVENTION

[0002]This invention is in the field of plant molecular biology. More specifically, this invention pertains to nucleic acid fragments encoding enzymes involved in amino acid biosynthesis in plants and seeds.

BACKGROUND OF THE INVENTION

[0003]Many vertebrates, including humans, lack the ability to manufacture a number of amino acids and therefore require these amino acids in their diet. These are called essential amino acids. Grain-derived foods or feed, however, are deficient in certain essential amino acids, such as lysine, the sulfur-containing amino acids methionine and cysteine, threonine and tryptophan. For example, in corn (Zea mays L.) lysine is the most limiting amino acid for the dietary requirements of many animals, and soybean (Glycine max L.) meal is used as an additive to corn-based animal feeds primarily as a lysine supplement. Often microbial-fermentation produced lysine is needed for such supplementation. Thus, an increase in lysine content of either corn or soybean would reduce or eliminate the need to supplement mixed grain feeds with lysine produced via fermentation.

[0004]Furthermore, in corn the sulfur amino acids are the third most limiting amino acids, after lysine and tryptophan, for the dietary requirements of many animals. Legume plants, however, while rich in lysine and tryptophan, have low sulfur-containing amino acid content. Therefore, the use of soybean meal to supplement corn in animal feed is not satisfactory. An increase in the sulfur amino acid content of either corn or soybean would improve the nutritional quality of the mixtures and reduce the need for further supplementation through addition of more expensive methionine.

[0005]One approach to increasing the nutritional quality of human foods and animal feed is to increase the production and accumulation of specific free amino acids via genetic engineering of the biosynthetic pathway of the essential amino acids. Biosynthetically, lysine, threonine, methionine, cysteine and isoleucine are all derived from aspartate. Regulation of the biosynthesis of each member of this family is interconnected (see FIG. 1). The organization of the pathway leading to biosynthesis of lysine, threonine, methionine, cysteine and isoleucine indicates that over-expression or reduction of expression of genes encoding, inter alia, aspartic semialdehyde dehydrogenase, homoserine kinase, diaminopimelate decarboxylase, cysteine synthase and cystathionine β-lyase in corn and soybean could be used to alter levels of these amino acids in human food and animal feed. However, few of the genes encoding enzymes that regulate this pathway in plants, especially corn and soybeans, are available. Accordingly, availability of nucleic acid sequences encoding all or a portion of these enzymes would facilitate development of nutritionally improved crop plants.

SUMMARY OF THE INVENTION

[0006]The present invention relates to isolated polynucleotides selected from the group consisting of SEQ ID NOs:1, 3, 5, 42, 44, 46, 48, 50, 8, 10, 12, 14, 16, 18, 53, 55, 21, 23, 25, 27, 58, 30, 61, 63, 33, 35, 37, 39, 67, 69, and 71.

[0007]The present invention concerns isolated polynucleotides comprising a nucleotide sequence selected from the group consisting of: (a) a nucleotide sequence encoding a polypeptide of at least 60 amino acids having at least 80% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:2, 4, 6, 43, 45, 47, 49, and 51; (b) a nucleotide sequence encoding a polypeptide of at least 60 amino acids having at least 95% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:9, 11, 13, 15, 17, 19, 54 and 56; (c) a nucleotide sequence encoding a polypeptide of at least 60 amino acids having at least 80% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:22, 24, 26, 28, and 59; (d) a nucleotide sequence encoding a polypeptide of at least 60 amino acids having at least 95% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:31, 62, and 64; and (e) a nucleotide sequence encoding a polypeptide of at least 60 amino acids having at least 85% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:34, 36, 38, 40, 68, 70, and 72. It is preferred that the identity be at least 85%, more preferably at least 90%, still more preferably at least 95%. This invention also relates to the isolated complement of such polynucleotides, wherein the complement and the polynucleotide consist of the same number of nucleotides, and the nucleotide sequences of the complement and the polynucleotide have 100% complementarity.

[0008]In a third embodiment nucleotide sequence of the isolated first polynucleotide is selected from SEQ ID NOs:1, 3, 5, 42, 44, 46, 48, 50, SEQ ID NOs:8, 10, 12, 14, 16, 18, 53 and 55, SEQ ID NOs:21, 23, 25, 27, and 58, SEQ ID NOs:30, 61, and 63, and SEQ ID NOs:33, 35, 37, 39, 67, 69, and 71.

[0009]In a fourth embodiment, this invention concerns an isolated polynucleotide encoding an aspartic semialdehyde dehydrogenase, a diaminopimelate decarboxylase, a homoserine kinase, a cysteine γ synthase or a cystathionine β-lyase.

[0010]In a fifth embodiment, this invention relates to a chimeric gene comprising the polynucleotide of the present invention.

[0011]In a sixth embodiment, the present invention concerns an isolated nucleic acid molecule that comprises at least 180 nucleotides and remains hybridized with the isolated polynucleotide of the present invention under a wash condition of 0.1×SSC, 0.1% SDS, and 65° C.

[0012]In a seventh embodiment, the invention also relates to a host cell comprising a chimeric gene of the present invention or an isolated polynucleotide of the present invention. The host cell may be eukaryotic, such as a yeast cell or a plant cell, or prokaryotic, such as a bacterial cell. The present invention may also relate to a virus comprising an isolated polynucleotide of the present invention or a chimeric gene of the present invention.

[0013]In an eighth embodiment, the invention concerns a transgenic plant comprising a polynucleotide of the present invention.

[0014]In a ninth embodiment, the invention relates to a method for transforming a cell by introducing into such cell the polynucleotide of the present invention, or a method of producing a transgenic plant by transforming a plant cell with the polynucleotide of the present invention and regenerating a plant from the transformed plant cell.

[0015]In a tenth embodiment, the invention concerns a method for producing a nucleotide fragment by selecting a nucleotide sequence comprised by a polynucleotide of the present invention and synthesizing a polynucleotide fragment containing the nucleotide sequence. It is understood that the nucleotide fragment may be produced in vitro or in vivo.

[0016]In an eleventh embodiment the invention concerns an isolated polypeptide comprising an amino acid sequence selected from the group consisting of: (a) a polypeptide of at least 60 amino acids and having a sequence identity of at least 80% based on the Clustal method of alignment when compared to an amino acid sequence selected from the group consisting of SEQ ID NOs:2, 4, 6, 43, 45, 47, 49, and 51; (b) a polypeptide of at least 60 amino acids having a sequence identity of at least 95% based on the Clustal method of alignment when compared to an amino acid sequence selected from the group consisting of SEQ ID NOs:9, 11, 13, 15, 17, 19, 54 and 56; (c) a polypeptide of at least 60 amino acids having a sequence identity of at least 80% based on the Clustal method of alignment when compared to an amino acid sequence selected from the group consisting of SEQ ID NOs:22, 24, 26, 28, and 59; (d) polypeptide of at least 60 amino acids having an identity of at least 95% based on the Clustal method of alignment when compared to an amino acid sequence selected from the group consisting of SEQ ID NOs:31, 62, and 64; and (e) a polypeptide of at least 60 amino acids having a sequence identity of at least 85% based on the Clustal method of alignment when compared to an amino acid sequence selected from the group consisting of SEQ ID NOs:34, 36, 38, 40, 68, 70, and 72. It is preferred that the identity be at least 85%, it is more preferred if the identity is at least 90%, it is preferable that the identity be at least 95%.

[0017]In a twelfth embodiment the invention relates to an isolated polypleptide selected from SEQ ID NOs:2, 4, 6, 43, 45, 47, 49, and 51, SEQ ID NOs:9, 11, 13, 15, 17, 19, 54 and 56, SEQ ID NOs:22, 24, 26, 28, and 59, SEQ ID NOs:31, 62, and 64, and SEQ ID NOs:34, 36, 38, 40, 68, 70, and 72.

[0018]In a thirteenth embodiment, this invention concerns an isolated polypeptide having aspartic semialdehyde dehydrogenase, diaminopimelate decarboxylase, homoserine kinase, cysteine γ synthase, or cystathionine β-lyase function.

[0019]In a fourteenth embodiment, this invention relates to a method of altering the level of expression of a plant biosynthetic enzyme in a host cell comprising: transforming a host cell with a chimeric gene of the present invention; and growing the transformed host cell under conditions that are suitable for expression of the chimeric gene.

[0020]A further embodiment of the instant invention is a method for evaluating a compound for its ability to inhibit the activity of a plant biosynthetic enzyme selected from the group consisting of aspartic semialdehyde dehydrogenase, diaminopimelate decarboxylase, homoserine kinase, cysteine γ synthase and cystathionine β-lyase, the method comprising the steps of: (a) transforming a host cell with a chimeric gene comprising a nucleic acid fragment encoding a plant biosynthetic enzyme selected from the group consisting of aspartic semialdehyde dehydrogenase, diaminopimelate decarboxylase, homoserine kinase, cysteine synthase and cystathionine β-lyase, operably linked to regulatory sequences; (b) growing the transformed host cell under conditions that are suitable for expression of the chimeric gene wherein expression of the chimeric gene results in production of the biosynthetic enzyme in the transformed host cell; (c) optionally purifying the biosynthetic enzyme expressed by the transformed host cell; (d) treating the biosynthetic enzyme with a compound to be tested; and (e) comparing the activity of the biosynthetic enzyme that has been treated with a test compound to the activity of an untreated biosynthetic enzyme.

BRIEF DESCRIPTION OF THE DRAWINGS AND SEQUENCE LISTINGS

[0021]The invention can be more fully understood from the following detailed description and the accompanying drawings and Sequence Listing which form a part of this application.

[0022]FIG. 1 depicts the biosynthetic pathway for the aspartate family of amino acids. The following abbreviations are used: AK=aspartokinase; ASADH=aspartic semialdehyde dehydrogenase; DHDPS=dihydrodipicolinate synthase; DHDPR=dihydrodipicolinate reductase; DAPEP=diaminopimelate epimerase; DAPDC=diaminopimelate decarboxylase; HDH=homoserine dehydrogenase; HK=homoserine kinase; TS=threonine synthase; TD=threonine deaminase; CγS=cystathionine γ-synthase; CβL=cystathionine β-lyase; MS=methionine synthase; CS=cysteine synthase; and SAMS=S-adenosylmethionine synthase.

[0023]FIGS. 2 through 6 show the amino acid sequence alignments between the known art sequences for aspartic semialdehyde dehydrogenase, diaminopimelate decarboxylase, homoserine kinase, cysteine γ synthase, and cystathione β-lyase with the sequences included in this application. Alignments were performed using the Clustal a logarithm described in Higgins and Sharp (1989) (CABIOS 5:151-153). Amino acids conserved among all sequences are indicated by an asterisk (*) above the alignment. Dashes are used by the program to maximize the alignment. A description of FIGS. 2 through 6 follows:

[0024]FIG. 2 shows a comparison of the aspartic semialdehyde dehydrogenase amino acid sequences from corn contig assembled from clones p0003.cgpha22r:fis, cpe1c.pk009.b24, p0016.ctscp83r, and p00075.cslab16r (SEQ ID NO:43), rice clone rlr48.pk0003.d12 (SEQ ID NO:2), the contig of 5' RACE PCR and rice clone rlr48.pk0003.d12 (SEQ ID NO:45), soybean clones sfl1.pk0122.f9 (SEQ ID NO:6), ses9c.pk001.a15:fis (SEQ ID NO:47), and sfl1.pk0122.f9:fis (SEQ ID NO:49), wheat clones wr1.pk0004.cl 1 (SEQ ID NO:4) and wdk1c.pk014.n5:fis (SEQ ID NO:51) with the Legionella pneumophila (NCBI General Identifier No. 2645882; SEQ ID NO:7) and the Aquifex aeolicus sequences (NCBI General Identifier No. 6225258; SEQ ID NO:52). FIG. 2A: positions 1 through 120; FIG. 2B: positions 121 through 240; FIG. 2c: positions 241 through 360; FIG. 2D: positions 361 through 392.

[0025]FIG. 3 shows a comparison of the diaminopimelate decarboxylase amino acid sequences derived from corn clones cen3n.pk0067.a3 (SEQ ID NO:9) and cr1n.pk0103.d8 (SEQ ID NO:11), rice clone rl0n.pk0013.b9 (SEQ ID NO:13), soybean clones sr1.pk0132.cl (SEQ ID NO:15), sdp3c.pk001.o15 (SEQ ID NO:19) and sdp3c.pk001.o15:fis (SEQ ID NO:54), wheat clones wlk1.pk0012.c2 (SEQ ID NO:17) and wlk1.pk0012.c2:fis (SEQ ID NO:56) with the Pseudomonas aeruginosa (NCBI General Identifier No. 118304; SEQ ID NO:20) and Arabidopsis thaliana sequences (NCBI General Identifier No. 9279586; SEQ ID NO:57). FIG. 3A: positions 1 through 120; FIG. 3B: positions 121 through 240; FIG. 3c: positions 241 through 360; FIG. 3D: positions 361 through 480; FIG. 3E: positions 481 through 535.

[0026]FIG. 4 shows a comparison of the homoserine kinase amino acid sequences derived from corn clone cr1n.pk0009.g4 (SEQ ID NO:22), rice clones rca1c.pk005.k3 (SEQ ID NO:24) and rca1c.pk005.k3:fis (SEQ ID NO:59), soybean clone ses8w.pk0020.b5 (SEQ ID NO:26), wheat clone wl1n.pk0065.f2 (SEQ ID NO:28) with the Methanococcus jannaschii (NCBI General Identifier No. 1591748; SEQ ID NO:29) and the Arabidopsis thaliana sequences (NCBI General Identifier No. 4927412; SEQ ID NO:60). FIG. 4A: positions 1 through 180; FIG. 4B: positions 181 though 360; FIG. 4C: positions 361 through 396.

[0027]FIG. 5 shows a comparison of the cysteine γ synthase amino acid sequences derived from the corn contig assembled from clones cco1n.pk083 j4, chp2.pk0016.b1, cpd1c.pk004.b20, cr1n.pk0083.c5, csi1.pk0003.g6, and p0126.cn1cb49r (SEQ ID NO:62), rice clone rls6.pk0068.b7:fis (SEQ ID NO:64), soybean clone se3.05h06 (SEQ ID NO:31) with the Citrullus lanatus sequence (NCBI General Identifier No. 540497; SEQ ID NO:32), the Spinacia oleracea sequence (NCBI General Identifier No. 540497; SEQ ID NO:65), and the Solanum tuberosum sequence (NCBI General Identifier No. 11131628; SEQ ID NO:66). FIG. 5A: positions 1 through 180; FIG. 5B: positions 181 through 360; FIG. 5C: positions 361 through 424.

[0028]FIG. 6 shows a comparison of the amino acid sequences of the cystathionine β-lyase derived from corn clone cen1.pk0061.d4 (SEQ ID NO:34), corn contig assembled from clones p0005.cbmei71r, p0014.ctuui39r, p0109.cdadg47r, and p0125.czaay16r (SEQ ID NO:68), rice clone rlr12.pk0026.g1 (SEQ ID NO:36), the contig of 5' PCR and rice clone rlr12.pk0026.g1:fis (SEQ ID NO:70), soybean clone sfl1.pk0012.c4 (SEQ ID NO:38), and wheat clones wr1.pk0091.g6 (SEQ ID NO:40) and wr1.pk0091.g6:fis (SEQ ID NO:72) with the Arabidopsis thaliana sequence (NCBI General Identifier No. 1708993; SEQ ID NO:41). FIG. 6A: positions 1 through 120; FIG. 6B: positions 121 through 240; FIG. 6c: positions 241 through 360; FIG. 6D: positions 361 through 483.

[0029]Table 1 lists the polypeptides that are described herein, the designation of the cDNA clones that comprise the nucleic acid fragments encoding polypeptides representing all or a substantial portion of these polypeptides, and the corresponding identifier (SEQ ID NO:) as used in the attached Sequence Listing. The sequence descriptions and Sequence Listing attached hereto comply with the rules governing nucleotide and/or amino acid sequence disclosures in patent applications as set forth in 37 C.F.R. §1.821-1.825.

TABLE-US-00001 TABLE 1 Plant Biosynthetic Enzymes SEQ ID NO: (Amino Polypeptide Clone (Nucleotide) Acid) rice ASADH rlr48.pk0003.d12 1 2 wheat ASADH wr1.pk0004.c11 3 4 soybean ASADH sfl1.pk0122.f9 5 6 L. pneumophila ASADH NCBI GI 2645882 7 corn DAPEP cen3n.pk0067.a3 8 9 corn DAPEP cr1n.pk0103.d8 10 11 rice DAPEP rl0n.pk0013.b9 12 13 soybean DAPEP sr1.pk0132.c1 14 15 wheat DAPEP wlk1.pk0012.c2 16 17 soybean DAPEP sdp3c.pk001.o15 18 19 P. aeruginosa DAPEP NCBI GI 118304 20 corn HK cr1n.pk0009.g4 21 22 rice HK rca1c.pk005.k3 23 24 soybean HK ses8w.pk0020.b5 25 26 wheat HK wl1n.pk0065.f2 27 28 M. jannaschii HK NCBI GI 1591748 29 soybean CγS se3.05h06 30 31 C. lanatus CγS NCBI GI 540497 32 corn CβL cen1.pk0061.d4 33 34 rice CβL rlr12.pk0026.g1 35 36 soybean CβL sfl1.pk0012.c4 37 38 wheat CβL wr1.pk0091.g6 39 40 A. thaliana CβL NCBI GI 1708993 41 corn ASADH Contig of: 42 43 p0003.cgpha22r:fis cpe1c.pk009.b24 p0016.ctscp83r p0075.cslab16r rice ASADH 5' RACE PCR + 44 45 rlr48.pk0003.d12 soybean ASADH ses9c.pk001.a15:fis 46 47 soybean ASADH sfl1.pk0122.f9:fis 48 49 wheat ASADH wdk1c.pk014.n5:fis 50 51 A. aeolicus ASADH NCBI GI 6225258 52 soybean DAPEP sdp3c.pk001.o15:fis 53 54 wheat DAPEP wlk1.pk0012.c2:fis 55 56 A. thaliana DAPEP NCBI GI 9279586 57 rice HK rca1c.pk005.k3:fis 58 59 A. thaliana HK NCBI GI 4927412 60 corn CγS Contig of: 61 62 cco1n.pk083.j4 chp2.pk0016.b1 cpd1c.pk004.b20 cr1n.pk0083.c5 csi1.pk0003.g6 p0126.cnlcb49r rice CγS rls6.pk0068.b7:fis 63 64 S. oleracea CγS NCBI GI 416869 65 S. tuberosum CγS NCBI GI 11131628 66 corn CβL Contig of: 67 68 p0005.cbmei71r p0014.ctuui39r p0109.cdadg47r p0125.czaay16r rice CβL 5'RACE PCR + 69 70 rlr12.pk0026.g1:fis wheat CβL wr1.pk0091.g6:fis 71 72

[0030]The nucleotide and amino acid sequences shown in SEQ ID NOs:1 through 41 are found, with the same SEQ ID NO, in U.S. application Ser. No. 09/424,976. All or a portion of some of the sequences in the present application are found in the provisional applications for which the present application claims priority to. Table 1A indicates the SEQ ID NO: in the present application and the corresponding SEQ ID NO: in the previously-filed provisional application.

TABLE-US-00002 TABLE 1A Sequence Priority Application Provisional Application Provisional Application No. 09/424,976 No. 60/049406 No. 60/065385 SEQ ID NO: 1 SEQ ID NO: 1 SEQ ID NO: 2 SEQ ID NO: 2 SEQ ID NO: 3 SEQ ID NO: 3* SEQ ID NO: 4 SEQ ID NO: 4* SEQ ID NO: 8 SEQ ID NO: 7 SEQ ID NO: 8 SEQ ID NO: 9 SEQ ID NO: 8 SEQ ID NO: 9 SEQ ID NO: 12 SEQ ID NO: 9 SEQ ID NO: 13 SEQ ID NO: 10 SEQ ID NO: 14 SEQ ID NO: 11 SEQ ID NO: 5 SEQ ID NO: 15 SEQ ID NO: 12 SEQ ID NO: 6 SEQ ID NO: 21 SEQ ID NO: 13 SEQ ID NO: 10* SEQ ID NO: 22 SEQ ID NO: 14 SEQ ID NOs: 11* and 14* SEQ ID NO: 23 SEQ ID NO: 17* SEQ ID NO: 15 SEQ ID NO: 24 SEQ ID NO: 18* SEQ ID NO: 16 SEQ ID NO: 25 SEQ ID NO: 15 SEQ ID NO: 13 SEQ ID NO: 26 SEQ ID NO: 16 SEQ ID NO: 14 SEQ ID NO: 30 SEQ ID NO: 19 SEQ ID NO: 17 SEQ ID NO: 31 SEQ ID NO: 20 SEQ ID NO: 18 SEQ ID NO: 33* SEQ ID NO: 21 SEQ ID NO: 19 SEQ ID NO: 34 SEQ ID NO: 22 SEQ ID NO: 20 SEQ ID NO: 37 SEQ ID NO: 23 SEQ ID NO: 21* SEQ ID NO: 38 SEQ ID NO: 24 SEQ ID NO: 22* *Indicates that only a portion of the sequence was in the application.

[0031]The Sequence Listing contains the one letter code for nucleotide sequence characters and the three letter codes for amino acids as defined in conformity with the IUPAC-IUBMB standards described in Nucleic Acids Res. 13:3021-3030 (1985) and in the Biochemical J. 219 (No. 2):345-373 (1984) which are herein incorporated by reference. The symbols and format used for nucleotide and amino acid sequence data comply with the rules set forth in 37 C.F.R. §1.822.

DETAILED DESCRIPTION OF THE INVENTION

[0032]In the context of this disclosure, a number of terms shall be utilized. The terms "polynucleotide," "polynucleotide sequence," "nucleic acid sequence," and "nucleic acid fragment"/"isolated nucleic acid fragment" are used interchangeably herein. These terms encompass nucleotide sequences and the like. A polynucleotide may be a polymer of RNA or DNA that is single- or double-stranded, that optionally contains synthetic, non-natural or altered nucleotide bases. A polynucleotide in the form of a polymer of DNA may be comprised of one or more segments of cDNA, genomic DNA, synthetic DNA, or mixtures thereof. An isolated polynucleotide of the present invention may include at least 30 contiguous nucleotides, preferably at least 40 contiguous nucleotides, most preferably at least 60 contiguous nucleotides derived from SEQ ID NOs:1, 3, 5, 42, 44, 46, 48, 50, SEQ ID NOs:8, 10, 12, 14, 16, 18, 53 and 55, SEQ ID NOs:21, 23, 25, 27, and 58, SEQ ID NOs:30, 61, and 63, and SEQ ID NOs:33, 35, 37, 39, 67, 69, and 71, or the complement of such sequences.

[0033]The term "isolated" polynucleotide refers to a polynucleotide that is substantially free from other nucleic acid sequences with which it is normally associated such as other chromosomal and extrachromosomal DNA and RNA. Isolated polynucleotides may be purified from a host cell in which they naturally occur. Conventional nucleic acid purification methods known to skilled artisans may be used to obtain isolated polynucleotides. The term also embraces recombinant polynucleotides and chemically synthesized polynucleotides.

[0034]The term "recombinant" means, for example, that a nucleic acid sequence is made by an artificial combination of two otherwise separated segments of sequence, e.g., by chemical synthesis or by the manipulation of isolated nucleic acids by genetic engineering techniques.

[0035]As used herein, "contig" refers to a nucleotide sequence that is assembled from two or more constituent nucleotide sequences that share common or overlapping regions of sequence homology. For example, the nucleotide sequences of two or more nucleic acid fragments can be compared and aligned in order to identify common or overlapping sequences. Where common or overlapping sequences exist between two or more nucleic acid fragments, the sequences (and thus their corresponding nucleic acid fragments) can be assembled into a single contiguous nucleotide sequence.

[0036]As used herein, "substantially similar" refers to nucleic acid fragments wherein changes in one or more nucleotide bases results in substitution of one or more amino acids, but do not affect the functional properties of the polypeptide encoded by the nucleotide sequence. "Substantially similar" also refers to nucleic acid fragments wherein changes in one or more nucleotide bases does not affect the ability of the nucleic acid fragment to mediate alteration of gene expression by gene silencing through for example antisense or co-suppression technology. "Substantially similar" also refers to modifications of the nucleic acid fragments of the instant invention such as deletion or insertion of one or more nucleotides that do not substantially affect the functional properties of the resulting transcript vis-a-vis the ability to mediate gene silencing or alteration of the functional properties of the resulting protein molecule. It is therefore understood that the invention encompasses more than the specific exemplary nucleotide or amino acid sequences and includes functional equivalents thereof. The terms "substantially similar" and "corresponding substantially" are used interchangeably herein.

[0037]Substantially similar nucleic acid fragments may be selected by screening nucleic acid fragments representing subfragments or modifications of the nucleic acid fragments of the instant invention, wherein one or more nucleotides are substituted, deleted and/or inserted, for their ability to affect the level of the polypeptide encoded by the unmodified nucleic acid fragment in a plant or plant cell. For example, a substantially similar nucleic acid fragment representing at least 30 contiguous nucleotides derived from the instant nucleic acid fragment can be constructed and introduced into a plant or plant cell. The level of the polypeptide encoded by the unmodified nucleic acid fragment present in a plant or plant cell exposed to the substantially similar nucleic fragment can then be compared to the level of the polypeptide in a plant or plant cell that is not exposed to the substantially similar nucleic acid fragment.

[0038]For example, it is well known in the art that antisense suppression and co-suppression of gene expression may be accomplished using nucleic acid fragments representing less than the entire coding region of a gene, and by using nucleic acid fragments that do not share 100% sequence identity with the gene to be suppressed. Moreover, alterations in a nucleic acid fragment which result in the production of a chemically equivalent amino acid at a given site, but do not affect the functional properties of the encoded polypeptide, are well known in the art. Thus, a codon for the amino acid alanine, a hydrophobic amino acid, may be substituted by a codon encoding another less hydrophobic residue, such as glycine, or a more hydrophobic residue, such as valine, leucine, or isoleucine. Similarly, changes which result in substitution of one negatively charged residue for another, such as aspartic acid for glutamic acid, or one positively charged residue for another, such as lysine for arginine, can also be expected to produce a functionally equivalent product. Nucleotide changes which result in alteration of the N-terminal and C-terminal portions of the polypeptide molecule would also not be expected to alter the activity of the polypeptide. Each of the proposed modifications is well within the routine skill in the art, as is determination of retention of biological activity of the encoded products. Consequently, an isolated polynucleotide comprising a nucleotide sequence of at least 30 (preferably at least 40, most preferably at least 60) contiguous nucleotides derived from a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 42, 44, 46, 48, 50, SEQ ID NOs:8, 10, 12, 14, 16, 18, 53 and 55, SEQ ID NOs:21, 23, 25, 27, and 58, SEQ ID NOs:30, 61, and 63, and SEQ ID NOs:33, 35, 37, 39, 67, 69, and 71 and the complement of such nucleotide sequences may be used in methods of selecting an isolated polynucleotide that affects the expression of an aspartic-semialdehyde dehydrogenase, a diaminopimelate decarboxylase, a homoserine kinase, a cysteine γ synthase, or a cystathionine β-lyase polypeptide in a host cell. A method of selecting an isolated polynucleotide that affects the level of expression of a polypeptide in a host cell may comprise the steps of: constructing an isolated polynucleotide of the present invention or an isolated chimeric gene of the present invention; introducing the isolated polynucleotide or the isolated chimeric gene into a host cell; measuring the level of a polypeptide or enzyme activity in the host cell containing the isolated polynucleotide; and comparing the level of a polypeptide or enzyme activity in the host cell containing the isolated polynucleotide with the level of a polypeptide or enzyme activity in a host cell that does not contain the isolated polynucleotide.

[0039]Moreover, substantially similar nucleic acid fragments may also be characterized by their ability to hybridize. Estimates of such homology are provided by either DNA-DNA or DNA-RNA hybridization under conditions of stringency as is well understood by those skilled in the art (Hames and Higgins, Eds. (1985) Nucleic Acid Hybridisation, IRL Press, Oxford, U.K.). Stringency conditions can be adjusted to screen for moderately similar fragments, such as homologous sequences from distantly related organisms, to highly similar fragments, such as genes that duplicate functional enzymes from closely related organisms. Post-hybridization washes determine stringency conditions. One set of preferred conditions uses a series of washes starting with 6×SSC, 0.5% SDS at room temperature for 15 min, then repeated with 2×SSC, 0.5% SDS at 45° C. for 30 min, and then repeated twice with 0.2×SSC, 0.5% SDS at 50° C. for 30 min. A more preferred set of stringent conditions uses higher temperatures in which the washes are identical to those above except for the temperature of the final two 30 min washes in 0.2×SSC, 0.5% SDS was increased to 60° C. Another preferred set of highly stringent conditions uses two final washes in 0.1 X SSC, 0.1% SDS at 65° C.

[0040]Substantially similar nucleic acid fragments of the instant invention may also be characterized by the percent identity of the amino acid sequences that they encode to the amino acid sequences disclosed herein, as determined by algorithms commonly employed by those skilled in this art. Suitable nucleic acid fragments (isolated polynucleotides of the present invention) encode polypeptides that are at least about 70% identical, preferably at least about 80% identical to the amino acid sequences reported herein. Preferred nucleic acid fragments encode amino acid sequences that are about 85% identical to the amino acid sequences reported herein. More preferred nucleic acid fragments encode amino acid sequences that are at least about 90% identical to the amino acid sequences reported herein. Most preferred are nucleic acid fragments that encode amino acid sequences that are at least about 95% identical to the amino acid sequences reported herein. Suitable nucleic acid fragments not only have the above identities but typically encode a polypeptide having at least 50 amino acids, preferably at least 100 amino acids, more preferably at least 150 amino acids, still more preferably at least 200 amino acids, and most preferably at least 250 amino acids. Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal method of alignment (Higgins and Sharp (1989) CABIOS. 5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5.

[0041]A "substantial portion" of an amino acid or nucleotide sequence comprises an amino acid or a nucleotide sequence that is sufficient to afford putative identification of the protein or gene that the amino acid or nucleotide sequence comprises. Amino acid and nucleotide sequences can be evaluated either manually, by one skilled in the art, or by using computer-based sequence comparison and identification tools that employ algorithms such as BLAST (Basic Local Alignment Search Tool; Altschul et al. (1993) J. Mol. Biol. 215:403-410; see also www.ncbi.nlm.nih.gov/BLAST/). In general, a sequence of ten or more contiguous amino acids or thirty or more contiguous nucleotides is necessary in order to putatively identify a polypeptide or nucleic acid sequence as homologous to a known protein or gene. Moreover, with respect to nucleotide sequences, gene-specific oligonucleotide probes comprising 30 or more contiguous nucleotides may be used in sequence-dependent methods of gene identification (e.g., Southern hybridization) and isolation (e.g., in situ hybridization of bacterial colonies or bacteriophage plaques). In addition, short oligonucleotides of 12 or more nucleotides may be used as amplification primers in PCR in order to obtain a particular nucleic acid fragment comprising the primers. Accordingly, a "substantial portion" of a nucleotide sequence comprises a nucleotide sequence that will afford specific identification and/or isolation of a nucleic acid fragment comprising the sequence. The instant specification teaches amino acid and nucleotide sequences encoding polypeptides that comprise one or more particular plant proteins. The skilled artisan, having the benefit of the sequences as reported herein, may now use all or a substantial portion of the disclosed sequences for purposes known to those skilled in this art. Accordingly, the instant invention comprises the complete sequences as reported in the accompanying Sequence Listing, as well as substantial portions of those sequences as defined above.

[0042]"Codon degeneracy" refers to divergence in the genetic code permitting variation of the nucleotide sequence without affecting the amino acid sequence of an encoded polypeptide. Accordingly, the instant invention relates to any nucleic acid fragment comprising a nucleotide sequence that encodes all or a substantial portion of the amino acid sequences set forth herein. The skilled artisan is well aware of the "codon-bias" exhibited by a specific host cell in usage of nucleotide codons to specify a given amino acid. Therefore, when synthesizing a nucleic acid fragment for improved expression in a host cell, it is desirable to design the nucleic acid fragment such that its frequency of codon usage approaches the frequency of preferred codon usage of the host cell.

[0043]"Synthetic nucleic acid fragments" can be assembled from oligonucleotide building blocks that are chemically synthesized using procedures known to those skilled in the art. These building blocks are ligated and annealed to form larger nucleic acid fragments which may then be enzymatically assembled to construct the entire desired nucleic acid fragment. "Chemically synthesized", as related to a nucleic acid fragment, means that the component nucleotides were assembled in vitro. Manual chemical synthesis of nucleic acid fragments may be accomplished using well established procedures, or automated chemical synthesis can be performed using one of a number of commercially available machines. Accordingly, the nucleic acid fragments can be tailored for optimal gene expression based on optimization of the nucleotide sequence to reflect the codon bias of the host cell. The skilled artisan appreciates the likelihood of successful gene expression if codon usage is biased towards those codons favored by the host. Determination of preferred codons can be based on a survey of genes derived from the host cell where sequence information is available.

[0044]"Gene" refers to a nucleic acid fragment that expresses a specific protein, including regulatory sequences preceding (5' non-coding sequences) and following (3' non-coding sequences) the coding sequence. "Native gene" refers to a gene as found in nature with its own regulatory sequences. "Chimeric gene" refers any gene that is not a native gene, comprising regulatory and coding sequences that are not found together in nature. Accordingly, a chimeric gene may comprise regulatory sequences and coding sequences that are derived from different sources, or regulatory sequences and coding sequences derived from the same source, but arranged in a manner different than that found in nature. "Endogenous gene" refers to a native gene in its natural location in the genome of an organism. A "foreign-gene" refers to a gene not normally found in the host organism, but that is introduced into the host organism by gene transfer. Foreign genes can comprise native genes inserted into a non-native organism, or chimeric genes. A "transgene" is a gene that has been introduced into the genome by a transformation procedure.

[0045]"Coding sequence" refers to a nucleotide sequence that codes for a specific amino acid sequence. "Regulatory sequences" refer to nucleotide sequences located upstream (5' non-coding sequences), within, or downstream (3' non-coding sequences) of a coding sequence, and which influence the transcription, RNA processing or stability, or translation of the associated coding sequence. Regulatory sequences may include promoters, translation leader sequences, introns, and polyadenylation recognition sequences.

[0046]"Promoter" refers to a nucleotide sequence capable of controlling the expression of a coding sequence or functional RNA. In general, a coding sequence is located 3' to a promoter sequence. The promoter sequence consists of proximal and more distal upstream elements, the latter elements often referred to as enhancers. Accordingly, an "enhancer" is a nucleotide sequence which can stimulate promoter activity and may be an innate element of the promoter or a heterologous element inserted to enhance the level or tissue-specificity of a promoter. Promoters may be derived in their entirety from a native gene, or may be composed of different elements derived from different promoters found in nature, or may even comprise synthetic nucleotide segments. It is understood by those skilled in the art that different promoters may direct the expression of a gene in different tissues or cell types, or at different stages of development, or in response to different environmental conditions. Promoters which cause a nucleic acid fragment to be expressed in most cell types at most times are commonly referred to as "constitutive promoters". New promoters of various types useful in plant cells are constantly being discovered; numerous examples may be found in the compilation by Okamuro and Goldberg (1989) Biochemistry of Plants 15: 1-82. It is further recognized that since in most cases the exact boundaries of regulatory sequences have not been completely defined, nucleic acid fragments of different lengths may have identical promoter activity.

[0047]"Translation leader sequence" refers to a nucleotide sequence located between the promoter sequence of a gene and the coding sequence. The translation leader sequence is present in the fully processed mRNA upstream of the translation start sequence. The translation leader sequence may affect processing of the primary transcript to mRNA, mRNA stability or translation efficiency. Examples of translation leader sequences have been described (Turner and Foster (1995) Mol. Biotechnol. 3:225-236).

[0048]"3' non-coding sequences" refer to nucleotide sequences located downstream of a coding sequence and include polyadenylation recognition sequences and other sequences encoding regulatory signals capable of affecting mRNA processing or gene expression. The polyadenylation signal is usually characterized by affecting the addition of polyadenylic acid tracts to the 3' end of the mRNA precursor. The use of different 3' non-coding sequences is exemplified by Ingelbrecht et al. (1989) Plant Cell 1:671-680.

[0049]"RNA transcript" refers to the product resulting from RNA polymerase-catalyzed transcription of a DNA sequence. When the RNA transcript is a perfect complementary copy of the DNA sequence, it is referred to as the primary transcript or it may be a RNA sequence derived from posttranscriptional processing of the primary transcript and is referred to as the mature RNA. "Messenger RNA (mRNA)" refers to the RNA that is without introns and that can be translated into polypeptides by the cell. "cDNA" refers to DNA that is complementary to and derived from an mRNA template. The cDNA can be single-stranded or converted to double stranded form using, for example, the Klenow fragment of DNA polymerase I. "Sense-RNA" refers to an RNA transcript that includes the mRNA and so can be translated into a polypeptide by the cell. "Antisense RNA" refers to an RNA transcript that is complementary to all or part of a target primary transcript or mRNA and that blocks the expression of a target gene (see U.S. Pat. No. 5,107,065, incorporated herein by reference). The complementarity of an antisense RNA may be with any part of the specific nucleotide sequence, i.e., at the 5' non-coding sequence, 3' non-coding sequence, introns, or the coding sequence. "Functional RNA" refers to sense RNA, antisense RNA, ribozyme RNA, or other RNA that may not be translated but yet has an effect on cellular processes.

[0050]The term "operably linked" refers to the association of two or more nucleic acid fragments on a single polynucleotide so that the function of one is affected by the other. For example, a promoter is operably linked with a coding sequence when it is capable of affecting the expression of that coding sequence (i.e., that the coding sequence is under the transcriptional control of the promoter). Coding sequences can be operably linked to regulatory sequences in sense or antisense orientation.

[0051]The term "expression", as used herein, refers to the transcription and stable accumulation of sense (mRNA) or antisense RNA derived from the nucleic acid fragment of the invention. Expression may also refer to translation of mRNA into a polypeptide. "Antisense inhibition" refers to the production of antisense RNA transcripts capable of suppressing the expression of the target protein. "Overexpression" refers to the production of a gene product in transgenic organisms that exceeds levels of production in normal or non-transformed organisms. "Co-suppression" refers to the production of sense RNA transcripts capable of suppressing the expression of identical or substantially similar foreign or endogenous genes (U.S. Pat. No. 5,231,020, incorporated herein by reference).

[0052]A "protein" or "polypeptide" is a chain of amino acids arranged in a specific order determined by the coding sequence in a polynucleotide encoding the polypeptide. Each protein or polypeptide has a unique function.

[0053]"Altered levels" or "altered expression" refers to the production of gene product(s) in transgenic organisms in amounts or proportions that differ from that of normal or non-transformed organisms.

[0054]"Mature protein" or the term "mature" when used in describing a protein refers to a post-translationally processed polypeptide; i.e., one from which any pre- or propeptides present in the primary translation product have been removed. "Precursor protein" or the term "precursor" when used in describing a protein refers to the primary product of translation of mRNA; i.e., with pre- and propeptides still present. Pre- and propeptides may be but are not limited to intracellular localization signals.

[0055]A "chloroplast transit peptide" is an amino acid sequence which is translated in conjunction with a protein and directs the protein to the chloroplast or other plastid types present in the cell in which the protein is made. "Chloroplast transit sequence" refers to a nucleotide sequence that encodes a chloroplast transit peptide. A "signal peptide" is an amino acid sequence which is translated in conjunction with a protein and directs the protein to the secretory system (Chrispeels (1991) Ann. Rev. Plant Phys. Plant Mol. Biol. 42:21-53). If the protein is to be directed to a vacuole, a vacuolar targeting signal (supra) can further be added, or if to the endoplasmic reticulum, an endoplasmic reticulum retention signal (supra) may be added. If the protein is to be directed to the nucleus, any signal peptide present should be removed and instead a nuclear localization signal included (Raikhel (1992) Plant Phys. 100:1627-1632).

[0056]"Transformation" refers to the transfer of a nucleic acid fragment into the genome of a host organism, resulting in genetically stable inheritance. Host organisms containing the transformed nucleic acid fragments are referred to as "transgenic" organisms. Examples of methods of plant transformation include Agrobacterium-mediated transformation (De Blaere et al. (1987) Meth. Enzymol. 143:277) and particle-accelerated or "gene gun" transformation technology (Klein et al. (1987) Nature (London) 327:70-73; U.S. Pat. No. 4,945,050, incorporated herein by reference). Thus, isolated polynucleotides of the present invention can be incorporated into recombinant constructs, typically DNA constructs, capable of introduction into and replication in a host cell. Such a construct can be a vector that includes a replication system and sequences that are capable of transcription and translation of a polypeptide-encoding sequence in a given host cell. A number of vectors suitable for stable transfection of plant cells or for the establishment of transgenic plants have been described in, e.g., Pouwels et al., Cloning Vectors: A Laboratory Manual, 1985, supp. 1987; Weissbach and Weissbach, Methods for Plant Molecular Biology, Academic Press, 1989; and Flevin et al., Plant Molecular Biology Manual, Kluwer Academic Publishers, 1990. Typically, plant expression vectors include, for example, one or more cloned plant genes under the transcriptional control of 5' and 3' regulatory sequences and a dominant selectable marker. Such plant expression vectors also can contain a promoter regulatory region (e.g., a regulatory region controlling inducible or constitutive, environmentally- or developmentally-regulated, or cell- or tissue-specific expression), a transcription initiation start site, a ribosome binding site, an RNA processing signal, a transcription termination site, and/or a polyadenylation signal.

[0057]Standard recombinant DNA and molecular cloning techniques used herein are well known in the art and are described more fully in Sambrook et al. Molecular Cloning: A Laboratory Manual; Cold Spring Harbor Laboratory Press: Cold Spring Harbor, 1989 (hereinafter "Maniatis").

[0058]"PCR" or "polymerase chain reaction" is well known by those skilled in the art as a technique used for the amplification of specific DNA segments (U.S. Pat. Nos. 4,683,195 and 4,800,159).

[0059]The present invention concerns isolated polynucleotides comprising a nucleotide sequence selected from the group consisting of: (a) a nucleotide sequence encoding a polypeptide of at least 60 amino acids having at least 80% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:2, 4, 6, 43, 45, 47, 49, and 51; (b) a nucleotide sequence encoding a polypeptide of at least 60 amino acids having at least 95% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:9, 11, 13, 15, 17, 19, 54 and 56; (c) a nucleotide sequence encoding a polypeptide of at least 60 amino acids having at least 80% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:22, 24, 26, 28, and 59; (d) a nucleotide sequence encoding a polypeptide of at least 60 amino acids having at least 95% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:31, 62, and 64; and (e) a nucleotide sequence encoding a polypeptide of at least 60 amino acids having at least 85% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:34, 36, 38, 40, 68, 70, and 72. It is preferred that the identity be at least 85%, it is preferable if the identity is at least 90%, it is more preferred that the identity be at least 95%. This invention also relates to the isolated complement of such polynucleotides, wherein the complement and the polynucleotide consist of the same number of nucleotides, and the nucleotide sequences of the complement and the polynucleotide have 100% complementarity.

[0060]Preferably, the isolated polynucleotide of the claimed invention comprises a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 42, 44, 46, 48, 50, 8, 10, 12, 14, 16, 18, 53, 55, 21, 23, 25, 27, 58, 30, 61, 63, 33, 35, 37, 39, 67, 69, and 71.

[0061]Nucleic acid fragments encoding at least a portion of several plant amino acid biosynthetic enzymes have been isolated and identified by comparison of random plant cDNA sequences to public databases containing nucleotide and protein sequences using the BLAST algorithms well known to those skilled in the art. The nucleic acid fragments of the instant invention may be used to isolate cDNAs and genes encoding homologous proteins from the same or other plant species. Isolation of homologous genes using sequence-dependent protocols is well known in the art. Examples of sequence-dependent protocols include, but are not limited to, methods of nucleic acid hybridization, and methods of DNA and RNA amplification as exemplified by various uses of nucleic acid amplification technologies (e.g., polymerase chain reaction, ligase chain reaction).

[0062]For example, genes encoding other aspartic semialdehyde dehydrogenases, diaminopimelate decarboxylases, homoserine kinases, cysteine γ synthases or cystathionine β-lyases, either as cDNAs or genomic DNAs, could be isolated directly by using all or a portion of the instant nucleic acid fragments as DNA hybridization probes to screen libraries from any desired plant employing methodology well known to those skilled in the art. Specific oligonucleotide probes based upon the instant nucleic acid sequences can be designed and synthesized by methods known in the art (Maniatis). Moreover, an entire sequence can be used directly to synthesize DNA probes by methods known to the skilled artisan such as random primer DNA labeling, nick translation, end-labeling techniques, or RNA probes using available in vitro transcription systems. In addition, specific primers can be designed and used to amplify a part or all of the instant sequences. The resulting amplification products can be labeled directly during amplification reactions or labeled after amplification reactions, and used as probes to isolate full length cDNA or genomic fragments under conditions of appropriate stringency.

[0063]In addition, two short segments of the instant nucleic acid fragments may be used in polymerase chain reaction protocols to amplify longer nucleic acid fragments encoding homologous genes from DNA or RNA. The polymerase chain reaction may also be performed on a library of cloned nucleic acid fragments wherein the sequence of one primer is derived from the instant nucleic acid fragments, and the sequence of the other primer takes advantage of the presence of the polyadenylic acid tracts to the 3' end of the mRNA precursor encoding plant genes. Alternatively, the second primer sequence may be based upon sequences derived from the cloning vector. For example, the skilled artisan can follow the RACE protocol (Frohman et al. (1988) Proc. Natl. Acad. Sci. USA 85:8998-9002) to generate cDNAs by using PCR to amplify copies of the region between a single point in the transcript and the 3' or 5' end. Primers oriented in the 3' and 5' directions can be designed from the instant sequences. Using commercially available 3' RACE or 5' RACE systems (BRL), specific 3' or 5' cDNA fragments can be isolated (Ohara et al. (1989) Proc. Natl. Acad. Sci. USA 86:5673-5677; Loh et al. (1989) Science 243:217-220). Products generated by the 3' and 5' RACE procedures can be combined to generate full-length cDNAs (Frohman and Martin (1989) Techniques 1:165). Consequently, a polynucleotide comprising a nucleotide sequence of at least 30 (preferably at least 40, most preferably at least 60) contiguous nucleotides derived from a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 42, 44, 46, 48, 50, 8, 10, 12, 14, 16, 18, 59, 61, 21, 23, 25, 27, 64, 30, 33, 35, 37, 39, 53, 55, and 57 and the complement of such nucleotide sequences may be used in such methods to obtain a nucleic acid fragment encoding a substantial portion of an amino acid sequence of a polypeptide.

[0064]The present invention relates to a method of obtaining a nucleic acid fragment encoding a substantial portion of an aspartic semialdehyde dehydrogenase, diaminopimelate decarboxylase, homoserine kinase, cysteine synthase, or cystathionine β-lyase polypeptide, preferably a substantial portion of a plant aspartic semialdehyde dehydrogenase, diaminopimelate decarboxylase, homoserine kinase, cysteine synthase, or cystathionine β-lyase polypeptide, comprising the steps of: synthesizing an oligonucleotide primer comprising a nucleotide sequence of at least 30 (preferably at least 40, most preferably at least 60) contiguous nucleotides derived from a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 42, 44, 46, 48, 50, 8, 10, 12, 14, 16, 18, 53, 55, 21, 23, 25, 27, 58, 30, 61, 63, 33, 35, 37, 39, 67, 69, and 71, and the complement of such nucleotide sequences; and amplifying a nucleic acid fragment (preferably a cDNA inserted in a cloning vector) using the oligonucleotide primer. The amplified nucleic acid fragment preferably will encode a portion of an aspartic semialdehyde dehydrogenase, diaminopimelate decarboxylase, homoserine kinase, cysteine synthase, or cystathionine β-lyase polypeptide.

[0065]Availability of the instant nucleotide and deduced amino acid sequences facilitates immunological screening of cDNA expression libraries. Synthetic peptides representing portions of the instant amino acid sequences may be synthesized. These peptides can be used to immunize animals to produce polyclonal or monoclonal antibodies with specificity for peptides or proteins comprising the amino acid sequences. These antibodies can be then be used to screen cDNA expression libraries to isolate full-length cDNA clones of interest (Lerner (1984) Adv. Immunol. 36:1-34; Maniatis).

[0066]In another embodiment, this invention concerns viruses and host cells comprising either the chimeric genes of the invention as described herein or an isolated polynucleotide of the invention as described herein. Examples of host cells which can be used to practice the invention include, but are not limited to, yeast, bacteria, and plants.

[0067]As was noted above, the nucleic acid fragments of the instant invention may be used to create transgenic plants in which the disclosed polypeptides are present at higher or lower levels than normal or in cell types or developmental stages in which they are not normally found. This would have the effect of altering the level of free amino acids in those cells. Specifically, the enzymes of the present invention form part of the pathway towards the biosynthesis of lysine, threonine, methionine, cysteine and isoleucine. In particular, altering the level and/or function of cystathionine beta-lyase will result in changes in the rate of methionine biosynthesis. Altering the level and/or function of diaminopimelate decarboxylase will result in changes in the rate of lysine biosynthesis. Altering the level and/or function of aspartate-semialdehyde dehydrogenase will result in changes in the lysine, methionine, or threonine content, especially in wheat. Altering the level of cysteine γ synthase will result in changes in the rate of cysteine and/or methionine biosynthesis; using this gene it will also be possible to control sulfur metabolism. Altering the level of homoserine kinase may be used to regulate threonine and methionine levels. Polypeptides encoding at least a portion of aspartic semialdehyde dehydrogenase, diaminopimelate decarboxylase, homoserine kinase, cysteine synthase, or cystathionine β-lyase may also be used in herbicide identification and design.

[0068]Overexpression of the proteins of the instant invention may be accomplished by first constructing a chimeric gene in which the coding region is operably linked to a promoter capable of directing expression of a gene in the desired tissues at the desired stage of development. The chimeric gene may comprise promoter sequences and translation leader sequences derived from the same genes. 3' Non-coding sequences encoding transcription termination signals may also be provided. The instant chimeric gene may also comprise one or more introns in order to facilitate gene expression.

[0069]Plasmid vectors comprising the instant isolated polynucleotide (or chimeric gene) may be constructed. The choice of plasmid vector is dependent upon the method that will be used to transform host plants. The skilled artisan is well aware of the genetic elements that must be present on the plasmid vector in order to successfully transform, select and propagate host cells containing the chimeric gene. The skilled artisan will also recognize that different independent transformation events will result in different levels and patterns of expression (Jones et al. (1985) EMBO J. 4:2411-2418; De Almeida et al. (1989) Mol. Gen. Genetics 218:78-86), and thus that multiple events must be screened in order to obtain lines displaying the desired expression level and pattern. Such screening may be accomplished by Southern analysis of DNA, Northern analysis of mRNA expression, Western analysis of protein expression, or phenotypic analysis.

[0070]For some applications it may be useful to direct the instant polypeptides to different cellular compartments, or to facilitate its secretion from the cell. It is thus envisioned that the chimeric gene described above may be further supplemented by directing the coding sequence to encode the instant polypeptides with appropriate intracellular targeting sequences such as transit sequences (Keegstra (1989) Cell 56:247-253), signal sequences or sequences encoding endoplasmic reticulum localization (Chrispeels (1991) Ann. Rev. Plant Phys. Plant Mol. Biol. 42:21-53), or nuclear localization signals (Raikhel (1992) Plant Phys. 100:1627-1632) with or without removing targeting sequences that are already present. While the references cited give examples of each of these, the list is not exhaustive and more targeting signals of use may be discovered in the future.

[0071]It may also be desirable to reduce or eliminate expression of genes encoding the instant polypeptides in plants for some applications. In order to accomplish this, a chimeric gene designed for co-suppression of the instant polypeptide can be constructed by linking a gene or gene fragment encoding that polypeptide to plant promoter sequences. Alternatively, a chimeric gene designed to express antisense RNA for all or part of the instant nucleic acid fragment can be constructed by linking the gene or gene fragment in reverse orientation to plant promoter sequences. Either the co-suppression or antisense chimeric genes could be introduced into plants via transformation wherein expression of the corresponding endogenous genes are reduced or eliminated.

[0072]Molecular genetic solutions to the generation of plants with altered gene expression have a decided advantage over more traditional plant breeding approaches. Changes in plant phenotypes can be produced by specifically inhibiting expression of one or more genes by antisense inhibition or cosuppression (U.S. Pat. Nos. 5,190,931, 5,107,065 and 5,283,323). An antisense or cosuppression construct would act as a dominant negative regulator of gene activity. While conventional mutations can yield negative regulation of gene activity these effects are most likely recessive. The dominant negative regulation available with a transgenic approach may be advantageous from a breeding perspective. In addition, the ability to restrict the expression of a specific phenotype to the reproductive tissues of the plant by the use of tissue specific promoters may confer agronomic advantages relative to conventional mutations which may have an effect in all tissues in which a mutant gene is ordinarily expressed.

[0073]The person skilled in the art will know that special considerations are associated with the use of antisense or cosuppression technologies in order to reduce expression of particular genes. For example, the proper level of expression of sense or antisense genes may require the use of different chimeric genes utilizing different regulatory elements known to the skilled artisan. Once transgenic plants are obtained by one of the methods described above, it will be necessary to screen individual transgenics for those that most effectively display the desired phenotype. Accordingly, the skilled artisan will develop methods for screening large numbers of transformants. The nature of these screens will generally be chosen on practical grounds. For example, one can screen by looking for changes in gene expression by using antibodies specific for the protein encoded by the gene being suppressed, or one could establish assays that specifically measure enzyme activity. A preferred method will be one which allows large numbers of samples to be processed rapidly, since it will be expected that a large number of transformants will be negative for the desired phenotype.

[0074]In another embodiment, the present invention concerns an aspartic-semialdehyde dehydrogenase polypeptide of at least 50 amino acids comprising at least 70% identity based on the Clustal method of alignment compared to a polypeptide selected from the group consisting of SEQ ID NOs:2, 4, 6, 43, 45, 47, 49, and 51, a diaminopimelate decarboxylase polypeptide of at least 60 amino acids comprising at least 95% identity based on the Clustal method of alignment compared to a polypeptide selected from the group consisting of SEQ ID NOs:9, 11, 13, 15, 17, 19, 60, and 62, a homoserine kinase polypeptide of at least 60 amino acids comprising at least 70% identity based on the Clustal method of alignment compared to a polypeptide selected from the group consisting of SEQ ID NOs:22, 24, 26, 28, and 65, a cysteine synthase polypeptide of at least 60 amino acids comprising at least 90% identity based on the Clustal method of alignment compared to a polypeptide of SEQ ID NO:31, or a cystathionine β-lyase polypeptide of at least 60 amino acids comprising at least 85% identity based on the Clustal method of alignment compared to a polypeptide selected from the group consisting of SEQ ID NOs:34, 36, 38, 40, 54, 56, and 58.

[0075]The instant polypeptides (or portions thereof) may be produced in heterologous host cells, particularly in the cells of microbial hosts, and can be used to prepare antibodies to these proteins by methods well known to those skilled in the art. The antibodies are useful for detecting the polypeptides of the instant invention in situ in cells or in vitro in cell extracts. Preferred heterologous host cells for production of the instant polypeptides are microbial hosts. Microbial expression systems and expression vectors containing regulatory sequences that direct high level expression of foreign proteins are well known to those skilled in the art. Any of these could be used to construct a chimeric gene for production of the instant polypeptides. This chimeric gene could then be introduced into appropriate microorganisms via transformation to provide high level expression of the encoded plant biosynthetic enzymes. An example of a vector for high level expression of the instant polypeptides in a bacterial host is provided (Example 10).

[0076]Additionally, the instant polypeptides can be used as a target to facilitate design and/or identification of inhibitors of those enzymes that may be useful as herbicides. This is desirable because the polypeptides described herein catalyze various steps in a pathway leading to production of several essential amino acids. Accordingly, inhibition of the activity of one or more of the enzymes described herein could lead to inhibition of plant growth. Thus, the instant polypeptides could be appropriate for new herbicide discovery and design.

[0077]All or a substantial portion of the polynucleotides of the instant invention may also be used as probes for genetically and physically mapping the genes that they are a part of, and used as markers for traits linked to those genes. Such information may be useful in plant breeding in order to develop lines with desired phenotypes. For example, the instant nucleic acid fragments may be used as restriction fragment length polymorphism (RFLP) markers. Southern blots (Maniatis) of restriction-digested plant genomic DNA may be probed with the nucleic acid fragments of the instant invention. The resulting banding patterns may then be subjected to genetic analyses using computer programs such as MapMaker (Lander et al. (1987) Genomics 1:174-181) in order to construct a genetic map. In addition, the nucleic acid fragments of the instant invention may be used to probe Southern blots containing restriction endonuclease-treated genomic DNAs of a set of individuals representing parent and progeny of a defined genetic cross. Segregation of the DNA polymorphisms is noted and used to calculate the position of the instant nucleic acid sequence in the genetic map previously obtained using this population (Botstein et al. (1980) Am. J. Hum. Genet. 32:314-331).

[0078]The production and use of plant gene-derived probes for use in genetic mapping is described in Bernatzky and Tanksley (1986) Plant Mol. Biol. Reporter 4:37-41. Numerous publications describe genetic mapping of specific cDNA clones using the methodology outlined above or variations thereof. For example, F2 intercross populations, backcross populations, randomly mated populations, near isogenic lines, and other sets of individuals may be used for mapping. Such methodologies are well known to those skilled in the art.

[0079]Nucleic acid probes derived from the instant nucleic acid sequences may also be used for physical mapping (i.e., placement of sequences on physical maps; see Hoheisel et al. In: Nonmammalian Genomic Analysis: A Practical Guide, Academic press 1996, pp. 319-346, and references cited therein).

[0080]In another embodiment, nucleic acid probes derived from the instant nucleic acid sequences may be used in direct fluorescence in situ hybridization (FISH) mapping (Trask (1991) Trends Genet. 7:149-154). Although current methods of FISH mapping favor use of large clones (several to several hundred KB; see Laan et al. (1995) Genome Res. 5:13-20), improvements in sensitivity may allow performance of FISH mapping using shorter probes.

[0081]A variety of nucleic acid amplification-based methods of genetic and physical mapping may be carried out using the instant nucleic acid sequences. Examples include allele-specific amplification (Kazazian (1989) J. Lab. Clin. Med. 11:95-96), polymorphism of PCR-amplified fragments (CAPS; Sheffield et al. (1993) Genomics 16:325-332), allele-specific ligation (Landegren et al. (1988) Science 241:1077-1080), nucleotide extension reactions (Sokolov (1990) Nucleic Acid Res. 18:3671), Radiation Hybrid Mapping (Walter et al. (1997) Nat. Genet. 7:22-28) and Happy Mapping (Dear and Cook (1989) Nucleic Acid Res. 17:6795-6807). For these methods, the sequence of a nucleic acid fragment is used to design and produce primer pairs for use in the amplification reaction or in primer extension reactions. The design of such primers is well known to those skilled in the art. In methods employing PCR-based genetic mapping, it may be necessary to identify DNA sequence differences between the parents of the mapping cross in the region corresponding to the instant nucleic acid sequence. This, however, is generally not necessary for mapping methods.

[0082]Loss of function mutant phenotypes may be identified for the instant cDNA clones either by targeted gene disruption protocols or by identifying specific mutants for these genes contained in a maize population carrying mutations in all possible genes (Ballinger and Benzer (1989) Proc. Natl. Acad. Sci. USA 86:9402-9406; Koes et al. (1995) Proc. Natl. Acad. Sci. USA 92:8149-8153; Bensen et al. (1995) Plant Cell 7:75-84). The latter approach may be accomplished in two ways. First, short segments of the instant nucleic acid fragments may be used in polymerase chain reaction protocols in conjunction with a mutation tag sequence primer on DNAs prepared from a population of plants in which Mutator transposons or some other mutation-causing DNA element has been introduced (see Bensen, supra). The amplification of a specific DNA fragment with these primers indicates the insertion of the mutation tag element in or near the plant gene encoding the instant polypeptides. Alternatively, the instant nucleic acid fragment may be used as a hybridization probe against PCR amplification products generated from the mutation population using the mutation tag sequence primer in conjunction with an arbitrary genomic site primer, such as that for a restriction enzyme site-anchored synthetic adaptor. With either method, a plant containing a mutation in the endogenous gene encoding the instant polypeptides can be identified and obtained. This mutant plant can then be used to determine or confirm the natural function of the instant polypeptides disclosed herein.

EXAMPLES

[0083]The present invention is further defined in the following Examples, in which parts and percentages are by weight and degrees are Celsius, unless otherwise stated. It should be understood that these Examples, while indicating preferred embodiments of the invention, are given by way of illustration only. From the above discussion and these Examples, one skilled in the art can ascertain the essential characteristics of this invention, and without departing from the spirit and scope thereof, can make various changes and modifications of the invention to adapt it to various usages and conditions. Thus, various modifications of the invention in addition to those shown and described herein will be apparent to those skilled in the art from the foregoing description. Such modifications are also intended to fall within the scope of the appended claims.

[0084]The disclosure of each reference set forth herein is incorporated herein by reference in its entirety.

Example 1

Composition of cDNA Libraries

Isolation and Sequencing of cDNA Clones

[0085]cDNA libraries representing mRNAs from various corn, rice, soybean, and wheat tissues were prepared. The characteristics of the libraries are described below.

TABLE-US-00003 TABLE 2 cDNA Libraries from Corn, Rice, Soybean, and Wheat Library Tissue Clone cen1 Corn Endosperm 12 Days After Pollination cen1.pk0061.d4 cen3n Corn Endosperm 20 Days After Pollination* cen3n.pk0067.a3 cpe1c Corn pooled BMS treated with chemicals related to cpe1c.pk009.b24 phosphatase** cr1n Corn Root From 7 Day Seedlings* cr1n.pk0009.g4 cr1n Corn Root From 7 Day Seedlings* cr1n.pk0103.d8 p0003 Corn Premeiotic Ear Shoot, 0.2-4 cm p0003.cgpha22r:fis p0005 Corn Immature Ear p0005.cbmei71r p0014 Corn Leaves 7 and 8 from Plant Transformed with p0014.ctuui39r G-protein Gene, C. heterostrophus Resistant p0016 Corn Tassel Shoots (0.1-1.4 cm), Pooled p0016.ctscp83r p0075 Corn Shoot And Leaf Material From p0075.cslab16r Dark-Grown 7 Day-Old Seedlings p0109 Corn Leaves From Les9 Transition Zone and Les9 Mature p0109.cdadg47r Lesions, Pooled*** p0125 Corn Anther Prophase 1* p0125.czaay16r rca1c Rice Nipponbare Callus rca1c.pk005.k3 rl0n Rice Leaf 15 Days After Germination* rl0n.pk0013.b9 rlr12 Rice Leaf 15 Days After Germination, 12 Hours After rlr12.pk0026.g1 Infection of Strain Magaporthe grisea 4360-R-62 (AVR2-YAMO) rlr48 Rice Leaf 15 Days After Germination 48 Hours After rlr48.pk0003.d12 Infection of Strain Magaporthe grisea 4360-R-62 (AVR2-YAMO) se3 Soybean Embryo 13 Days After Flowering sdp3c.pk001.o15 sdp3c Soybean Developing Pods 8-9 mm se3.05h06 ses8w Mature Soybean Embryo 8 Weeks After Subculture ses8w.pk0020.b5 ses9c Soybean Embryogenic Suspension ses9c.pk001.a15:fis sfl1 Soybean Immature Flower sfl1.pk0012.c4 sfl1 Soybean Immature Flower sfl1.pk0122.f9 sr1 Soybean Root From 10 Day Old Seedlings sr1.pk0132.c1 wdk1c Wheat Developing Kernel, 3 Days After Anthesis wdk1c.pk014.n5:fis wl1n Wheat Leaf from 7 Day Old Seedling* wl1n.pk0065.f2 wlk1 Wheat Seedlings 1 hour After Fungicide Treatment**** wlk1.pk0012.c2 wr1 Wheat Root From 7 Day Old Seedlings wr1.pk0004.c11 wr1 Wheat Root From 7 Day Old Seedlings wr1.pk0091.g6 *These libraries were normalized essentially as described in U.S. Pat. No. 5,482,845. **Chemicals used included okadaic acid, cyclosporin A, calyculin A, and cypermethrin, all of which are commercially available from Molecular Biology supply sources including Calbiochem-Novabiochem Corp. ***Les9 mutants reviewed in "An update on lesion mutants" Hoisington (1986) Maize Genetic Coop. News Lett. 60: 50-51. ****Application of 6-iodo-2-propoxy-3-propyl-4(3H)-quinazolinone; synthesis and methods of using this compound are described in USSN 08/545,827, incorporated herein by reference.

[0086]cDNA libraries may be prepared by any one of many methods available. For example, the cDNAs may be introduced into plasmid vectors by first preparing the cDNA libraries in Uni-ZAP® XR vectors according to the manufacturer's protocol (Stratagene Cloning Systems, La Jolla, Calif.). The Uni-ZAP® XR libraries are converted into plasmid libraries according to the protocol provided by Stratagene. Upon conversion, cDNA inserts will be contained in the plasmid vector pBluescript. In addition, the cDNAs may be introduced directly into precut Bluescript II SK(+) vectors (Stratagene) using T4 DNA ligase (New England Biolabs), followed by transfection into DH10B cells according to the manufacturer's protocol (GIBCO BRL Products). Once the cDNA inserts are in plasmid vectors, plasmid DNAs are prepared from randomly picked bacterial colonies containing recombinant pBluescript plasmids, or the insert cDNA sequences are amplified via polymerase chain reaction using primers specific for vector sequences flanking the inserted cDNA sequences. Amplified insert DNAs or plasmid DNAs are sequenced in dye-primer sequencing reactions to generate partial cDNA sequences (expressed sequence tags or "ESTs"; see Adams et al., (1991) Science 252:1651-1656). The resulting ESTs are analyzed using a Perkin Elmer Model 377 fluorescent sequencer.

[0087]Full-insert sequence (FIS) data is generated utilizing a modified transposition protocol. Clones identified for FIS are recovered from archived glycerol stocks as single colonies, and plasmid DNAs are isolated via alkaline lysis. Isolated DNA templates are reacted with vector primed M13 forward and reverse oligonucleotides in a PCR-based sequencing reaction and loaded onto automated sequencers. Confirmation of clone identification is performed by sequence alignment to the original EST sequence from which the FIS request is made.

[0088]Confirmed templates are transposed via the Primer Island transposition kit (PE Applied Biosystems, Foster City, Calif.) which is based upon the Saccharomyces cerevisiae Ty1 transposable element (Devine and Boeke (1994) Nucleic Acids Res. 22:3765-3772). The in vitro transposition system places unique binding sites randomly throughout a population of large DNA molecules. The transposed DNA is then used to transform DH10B electro-competent cells (Gibco BRL/Life Technologies, Rockville, Md.) via electroporation. The transposable element contains an additional selectable marker (named DHFR; Fling and Richards (1983) Nucleic Acids Res. 11:5147-5158), allowing for dual selection on agar plates of only those subclones containing the integrated transposon. Multiple subclones are randomly selected from each transposition reaction, plasmid DNAs are prepared via alkaline lysis, and templates are sequenced (ABI Prism dye-terminator ReadyReaction mix) outward from the transposition event site, utilizing unique primers specific to the binding sites within the transposon.

[0089]Sequence data is collected (ABI Prism Collections) and assembled using Phred/Phrap (P. Green, University of Washington, Seattle). Phrep/Phrap is a public domain software program which re-reads the ABI sequence data, re-calls the bases, assigns quality values, and writes the base calls and quality values into editable output files. The Phrap sequence assembly program uses these quality values to increase the accuracy of the assembled sequence contigs. Assemblies are viewed by the Consed sequence editor (D. Gordon, University of Washington, Seattle).

Example 2

Identification of cDNA Clones

[0090]cDNA clones encoding plant amino acid biosynthetic enzymes were identified by conducting BLAST (Basic Local Alignment Search Tool; Altschul et al. (1993) J. Mol. Biol. 215:403-410; see also www.ncbi.nlm.nih.gov/BLAST/) searches for similarity to sequences contained in the BLAST "nr" database (comprising all non-redundant GenBank CDS translations, sequences derived from the 3-dimensional structure Brookhaven Protein Data Bank, the last major release of the SWISS-PROT protein sequence database, EMBL, and DDBJ databases). The cDNA sequences obtained in Example 1 were analyzed for similarity to all publicly available DNA sequences contained in the "nr" database using the BLASTN algorithm provided by the National Center for Biotechnology Information (NCBI). The DNA sequences were translated in all reading frames and compared for similarity to all publicly available protein sequences contained in the "nr" database using the BLASTX algorithm (Gish and States (1993) Nat. Genet. 3:266-272) provided by the NCBI. For convenience, the P-value (probability) of observing a match of a cDNA sequence to a sequence contained in the searched databases merely by chance as calculated by BLAST are reported herein as "pLog" values, which represent the negative of the logarithm of the reported P-value. Accordingly, the greater the pLog value, the greater the likelihood that the cDNA sequence and the BLAST "hit" represent homologous proteins.

[0091]ESTs submitted for analysis are compared to the genbank database as described above. ESTs that contain sequences more 5- or 3-prime can be found by using the BLASTn algorithm (Altschul et al (1997) Nucleic Acids Res. 25:3389-3402.) against the DuPont proprietary database comparing nucleotide sequences that share common or overlapping regions of sequence homology. Where common or overlapping sequences exist between two or more nucleic acid fragments, the sequences can be assembled into a single contiguous nucleotide sequence, thus extending the original fragment in either the 5 or 3 prime direction. Once the most 5-prime EST is identified, its complete sequence can be determined by Full Insert Sequencing as described in Example 1. Homologous genes belonging to different species can be found by comparing the amino acid sequence of a known gene (from either a proprietary source or a public database) against an EST database using the tBLASTn algorithm. The tBLASTn algorithm searches an amino acid query against a nucleotide database that is translated in all 6 reading frames. This search allows for differences in nucleotide codon usage between different species, and for codon degeneracy.

Example 3

Characterization of cDNA Clones Encoding

Aspartate Semialdehyde Dehydrogenase

[0092]The BLASTX search using the EST sequences from clones listed in Table 3 revealed similarity of the polypeptides encoded by the cDNAs to aspartate semialdehyde dehydrogenase from Synechocystis sp. (DDJB Accession No. D64006; NCBI General Identifier No. 1001379) or Legionella pneumophila (GenBank Accession No. AF034213; NCBI General Identifier No. 2645882). Shown in Table 3 are the BLAST results for individual ESTs ("EST"), or for the sequences of the entire cDNA inserts comprising the indicated cDNA clones ("FIS"):

TABLE-US-00004 TABLE 3 BLAST Results for Sequences Encoding Polypeptides Homologous to Aspartate Semialdehyde Dehydrogenase BLAST pLog Score Synechocystis sp. Legionella pneumophila Clone Status GI 1001379 GI 2645882 rlr48.pk0003.d12 FIS 51.00 36.00 wr1.pk0004.c11 EST 67.96 44.74 sfl1.pk0122.f9 EST 6.60

[0093]The sequence of the entire cDNA insert in clone sfl1.pk0122.f9 was determined, RACE PCR was used to obtain the 5' portion of the rice aspartate semialdehyde dehydrogenase, and further sequencing and searching of the DuPont proprietary database allowed the identification of a corn and other a soybean, and wheat clones encoding aspartate semialdehyde dehydrogenase. The BLASTX search using the EST sequences from clones listed in Table 4 revealed similarity of the polypeptides encoded by the cDNAs to aspartate semialdehyde dehydrogenase from Aquifex aeolicus (NCBI General Identifier No. 6225258). Shown in Table 4 are the BLAST results for the sequences of contigs assembled from two or more ESTs ("Contig"), or the sequences encoding the entire protein derived from either the entire cDNA inserts comprising the indicated cDNA clones or contigs assembled from 5' RACE PCR and the sequence of the entire cDNA insert in the indicated cDNA clone ("CGS"):

TABLE-US-00005 TABLE 4 BLAST Results for Sequences Encoding Polypeptides Homologous to Aspartate Semialdehyde Dehydrogenase BLAST pLog Score Clone Status Aquifex aeolicus GI 6225258 Contig of: Contig 78.70 cpe1c.pk009.b24 p0003.cgpha22r:fis p0016.ctscp83r p0075.cslab16r 5' RACE PCR + CGS 89.20 rlr48.pk0003.d12:fis ses9c.pk001.a15:fis CGS 87.40 sfl1.pk0122.f9:fis CGS 88.10 wdk1c.pk014.n5:fis CGS 91.50

[0094]FIG. 2 presents an alignment of the amino acid sequences set forth in SEQ ID NOs:2, 4, 6, 43, 45, 47, 49, and 51 with the Legionella pneumophila sequence (NCBI General Identifier No. 2645882; SEQ ID NO:7) and the Aquifex aeolicus sequence (NCBI General Identifier No. 6225258; SEQ ID NO:52). The data in Table 5 presents a calculation of the percent identity of the amino acid sequences set forth in SEQ ID NOs:2, 4, 6, 43, 45, 47, 49, and 51 with the Legionella pneumophila sequence (NCBI General Identifier No. 2645882; SEQ ID NO:7) and the Aquifex aeolicus sequence (NCBI General Identifier No. 6225258; SEQ ID NO:52).

TABLE-US-00006 TABLE 5 Percent Identity of Amino Acid Sequences Deduced From the Nucleotide Sequences of cDNA Clones Encoding Polypeptides Homologous to Aspartate Semialdehyde Dehydrogenase amino acid Percent Identity to Clone SEQ ID NO. 2645882 6225258 rlr48.pk0003.d12 2 42.1 45.6 wr1.pk0004.c11 4 42.3 44.8 sfl1.pk0122.f9 6 29.1 25.6 Contig of: 43 41.2 45.9 cpe1c.pk009.b24 p0003.cgpha22r:fis p0016.ctscp83r p0075.cslab16r 5' RACE PCR + 45 43.2 47.0 rlr48.pk0003.d12:fis ses9c.pk001.a15:fis 47 43.5 49.1 sfl1.pk0122.f9:fis 49 41.2 45.6 wdk1c.pk014.n5:fis 51 43.2 49.4

[0095]As seen in FIG. 2, the amino acid sequence shown in SEQ ID NO:2 is identical to amino acids 181 through 375 of SEQ ID NO:45; the sequence shown in SEQ ID NO:4 is identical to amino acids 173 through 374 of the sequence shown in SEQ ID NO:51; the sequence shown in SEQ ID NO:6 is identical to amino acids 1 through 86 of the sequence shown in SEQ ID NO:49; there are 5 amino acid differences between the sequences shown in SEQ ID NO:47 and SEQ ID NO:49; there are 18 amino acid differences between amino acids 89 through 375 of the sequence shown in SEQ ID NO:43 and the sequence shown in SEQ ID NO:45; and there are 15 differences between the amino acid sequences shown in SEQ ID NO:45 and in SEQ ID NO:51.

[0096]Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal method of alignment (Higgins and Sharp (1989)CABIOS. 5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5. Sequence alignments and BLAST scores and probabilities indicate that the nucleic acid fragments comprising the instant cDNA clones encode a substantial portion of a corn aspartate semialdehyde dehydrogenase, a substantial portion and an entire rice aspartate semialdehyde dehydrogenase, a portion and an entire wheat aspartate semialdehyde dehydrogenase, and a portion and an two entire soybean aspartate semialdehyde dehydrogenases.

Example 4

Characterization of cDNA Clones Encoding Diaminopimelate Decarboxylase

[0097]The BLASTX search using the EST sequences from clones listed in Table 6 revealed similarity of the polypeptides encoded by the cDNAs to diaminopimelate decarboxylase from Aquifex aeolicus (GenBank Accession No. AE000728 and NCBI General Identifier No. 2983642) and Pseudomonas aeruginosa (GenBank Accession No. M23174 and NCBI General Identifier No. 118304). Shown in Table 6 are the BLAST results for individual ESTs ("EST"), the sequences of the entire cDNA inserts comprising the indicated cDNA clones ("FIS"), or the sequences of FISs encoding an entire protein ("CGS"):

TABLE-US-00007 TABLE 6 BLAST Results for Sequences Encoding Polypeptides Homologous to Diaminopimelate Decarboxylase BLAST pLog Score GI 2983642 GI 118304 Clone Status (A. aeolicus) (P. aeruginosa) cen3n.pk0067.a3 FIS 58.22 56.00 cr1n.pk0103.d8 CGS 75.25 79.12 rl0n.pk0013.b9 FIS 46.40 44.00 sr1.pk0132.c1 FIS 44.70 39.15 wlk1.pk0012.c2 EST 20.48 19.05

[0098]An additional soybean clone, sdp3c.pk001.o15, was identified as sharing homology with sr1.pk0132.cl. BLASTX search using the nucleotide sequences from clone sdp3c.pk001.o15 revealed similarity of the proteins encoded by the cDNA to diaminopimelate decarboxylase from Pseudomonas fluorescens (EMBO Accession No. Y12268; NCBI General Identifier No. 1929095). This EST yields a pLog value of 8.66 versus the Pseudomonas fluorescens sequence.

[0099]The sequence of the entire cDNA insert in clones sdp3c.pk001.o15 and wlk1.pk0012.c2 was determined. The BLASTX search using the EST sequences from clones listed in Table 7 revealed similarity of the polypeptides encoded by the cDNAs to diaminopimelate decarboxylase from Aquifex aeolicus (NCBI General Identifier No. 6225241) or by the Arabidopsis thaliana contig containing similarity with diaminopimelate decarboxylases (NCBI General Identifier No. 9279586). Shown in Table 7 are the BLAST results for the sequences of the entire cDNA inserts comprising the indicated cDNA clones ("FIS"), or the sequences of FISs encoding the entire protein ("CGS"):

TABLE-US-00008 TABLE 7 BLAST Results for Sequences Encoding Polypeptides Homologous to Diaminopimelate Decarboxylase BLAST Clone Status Homolog pLog Score sdp3c.pk001.o15:fis CGS GI 6225241 (A. aeolicus) 76.40 wlk1.pk0012.c2:fis FIS GI 9279586 (A. thaliana) 94.40

[0100]FIG. 3 presents an alignment of the amino acid sequences set forth in SEQ ID NOs:9, 11, 13, 15, 17, 19, 54, and 56 with the Pseudomonas aeruginosa sequence (NCBI General Identifier No. 118304; SEQ ID NO:20) and the Arabidopsis thaliana sequence (NCBI General Identifier No. 9279586, SEQ ID NO:57). The data in Table 8 presents a calculation of the percent identity of the amino acid sequences set forth in SEQ ID NOs:9, 11, 13, 15, 17, 19, 54, and 56 with the Pseudomonas aeruginosa sequence (NCBI General Identifier No. 118304; SEQ ID NO:20) and the Arabidopsis thaliana sequence (NCBI General Identifier No. 9279586; SEQ ID NO:57).

TABLE-US-00009 TABLE 8 Percent Identity of Amino Acid Sequences Deduced From the Nucleotide Sequences of cDNA Clones Encoding Polypeptides Homologous to Diaminopimelate Decarboxylase Amino acid Percent Identity to Clone SEQ ID NO. 118304 9279586 cen3n.pk0067.a3 9 34.0 82.2 cr1n.pk0103.d8 11 35.9 70.6 rl0n.pk0013.b9 13 32.4 76.8 sr1.pk0132.c1 15 29.7 86.1 wlk1.pk0012.c2 17 42.5 93.2 sdp3c.pk001.o15 19 41.9 87.1 sdp3c.pk001.o15:fis 54 32.5 74.9 wlk1.pk0012.c2:fis 56 32. 84.9

[0101]The amino acid sequence set forth in SEQ ID NO:19 is identical to amino acids 112 through 173 of the amino acid sequence set forth in SEQ ID NO:54. The amino acid sequence set forth in SEQ ID NO:17 is identical to amino acids 24 through 96 of the amino acid sequence set forth in SEQ ID NO:56.

[0102]Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal method of alignment (Higgins and Sharp (1989)CABIOS. 5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5. Sequence alignments and BLAST scores and probabilities indicate that the nucleic acid fragments comprising the instant cDNA clones encode a substantial portion of one corn, one rice, two soybean and one wheat diaminopimelate decarboxylases and entire corn and soybean diaminopimelate decarboxylases.

Example 5

Characterization of cDNA Clones Encoding Homoserine Kinase

[0103]The BLASTX search using the EST sequences from clones listed in Table 9 revealed similarity of the polypeptides encoded by the cDNAs to homoserine kinase from Methanococcus jannaschii (GenBank Accession No. U67553 and NCBI General Identifier No. 1591748). Shown in Table 9 are the BLAST results for individual ESTs ("EST") or for the sequences of the entire cDNA inserts comprising the indicated cDNA clones ("FIS"):

TABLE-US-00010 TABLE 9 BLAST Results for Sequences Encoding Polypeptides Homologous to Homoserine Kinase BLAST pLog Score Clone Status GI 1591748 (Methanococcus jannaschii) cr1n.pk0009.g4 FIS 19.30 rca1c.pk005.k3 EST 15.21 ses8w.pk0020.b5 FIS 35.30 wl1n.pk0065.f2 EST 5.68

[0104]The sequence of the entire cDNA insert in clone rca1c.pk005.k3 was determined. The BLASTX search using the EST sequences from clones listed in Table 10 revealed similarity of the polypeptides encoded by the cDNAs to homoserine kinase from Arabidopsis thaliana (NCBI General Identifier No. 4927412). Shown in Table 10 are the BLAST results for the sequences of the entire cDNA inserts comprising the indicated cDNA clone ("FIS"):

TABLE-US-00011 TABLE 10 BLAST Results for Sequences Encoding Polypeptides Homologous to Homoserine Kinase BLAST pLog Score Clone Status 4927412 (Arabidopsis thaliana) rca1c.pk005.k3:fis FIS 88.40

[0105]FIG. 4 presents an alignment of the amino acid sequences set forth in SEQ ID NOs:22, 24, 26, 28, and 59 with the Methanococcus jannaschii sequence (NCBI General Identifier No. 1591748; SEQ ID NO:29) and the Arabidopsis thaliana sequence (NCBI General Identifier No. 4927412; SEQ ID NO:60). The data in Table 11 presents a calculation of the percent identity of the amino acid sequences set forth in SEQ ID NOs:22, 24, 26, 28, and 59 with the Methanococcus jannaschii sequence (NCBI General Identifier No. 1591748; SEQ ID NO:29) and the Arabidopsis thaliana sequence (NCBI General Identifier No. 4927412; SEQ ID NO:60).

TABLE-US-00012 TABLE 11 Percent Identity of Amino Acid Sequences Deduced From the Nucleotide Sequences of cDNA Clones Encoding Polypeptides Homologous to Homoserine Kinase SEQ ID Percent Identity to Clone NO. NCBI GI 1591748 NCBI GI 4927412 cr1n.pk0009.g4 22 25.1 65.4 rca1c.pk005.k3 24 48.8 67.1 ses8w.pk0020.b5 26 28.0 65.7 wl1n.pk0065.f2 28 29.8 67.9 rca1c.pk005.k3:fis 59 28.6 65.9

[0106]The amino acid sequence set forth in SEQ ID NO:24 is identical to amino acids 18 through 99 of the amino acid sequence set forth in SEQ ID NO:59.

[0107]Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal method of alignment (Higgins and Sharp (1989)CABIOS. 5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5. Sequence alignments and BLAST scores and probabilities indicate that the nucleic acid fragments comprising the instant cDNA clones encode a substantial portion of a corn and a wheat homoserine kinase, a portion and an entire rice homoserine kinase, and an entire soybean homoserine kinase.

Example 6

Characterization of cDNA Clones Encoding Cysteine Synthase

[0108]The BLASTX search using the EST sequences from the clone listed in Table 12 revealed similarity of the polypeptides encoded by the cDNAs to cysteine synthase from Citrullus lanatus (DDJB Accession No. D28777, NCBI General Identifier No. 540497). Shown in Table 12 are the BLAST results for the sequences of the entire cDNA inserts comprising the indicated cDNA clones encoding the entire protein ("CGS"):

TABLE-US-00013 TABLE 12 BLAST Results for Sequences Encoding Polypeptides Homologous to Cysteine γ Synthase BLAST pLog Score Clone Status NCBI GI 540497 (Citrullus lanatus) se3.05h06 CGS 182.64

[0109]Further sequencing and searching of the DuPont proprietary database allowed the identification of corn and rice clones encoding polypeptides with similarities to cysteine γ synthase. The BLAST search using the sequences from clones listed in Table 13 revealed similarity of the polypeptides encoded by the cDNAs to cysteine γ synthase from Spinacia oleracea (NCBI General Identifier No. 416869) and Solanum tuberosum (NCBI General Identifier No. 11131628). Shown in Table 13 are the BLAST results for the sequences of the entire cDNA inserts comprising the indicated cDNA clones encoding the entire protein ("CGS"):

TABLE-US-00014 TABLE 13 BLAST Results for Sequences Encoding Polypeptides Homologous to Cysteine γ Synthase BLAST pLog Score NCBI GI 416869 NCBI GI 11131628 Clone Status (Spinacia oleracea) (Solanum tuberosum) Contig of: CGS 158.00 157.00 cco1n.pk083.j4 chp2.pk0016.b1 cpd1c.pk004.b20 cr1n.pk0083.c5 csi1.pk0003.g6 p0126.cnlcb49r rls6.pk0068.b7:fis CGS 161.00 163.00

[0110]FIG. 5 presents an alignment of the amino acid sequences set forth in SEQ ID NOs:31, 62, and 64 with the Citrullus lanatus sequence (NCBI General Identifier No. 540497; SEQ ID NO:32), Spinacia oleracea (NCBI General Identifier No. 416869; SEQ ID NO:65), and the Solanum tuberosum sequence (NCBI General Identifier No. 11131628; SEQ ID NO:66). The data in Table 14 presents a calculation of the percent identity of the amino acid sequences set forth in SEQ ID NOs:31, 62, and 64 with the Citrullus lanatus sequence (NCBI General Identifier No. 540497; SEQ ID NO:32), Spinacia oleracea (NCBI General Identifier No. 416869; SEQ ID NO:65), and the Solanum tuberosum sequence (NCBI General Identifier No. 11131628; SEQ ID NO:66).

TABLE-US-00015 TABLE 14 Percent Identity of Amino Acid Sequences Deduced From the Nucleotide Sequences of cDNA Clones Encoding Polypeptides Homologous to Cysteine γ Synthase Percent Identity to Amino acid NCBI NCBI NCBI Clone SEQ ID NO. GI 540497 GI 416869 GI 11131628 se3.05h06 31 87.1 72.3 76.9 Contig of: 62 73.8 71.3 69.7 cco1n.pk083.j4 chp2.pk0016.b1 cpd1c.pk004.b20 cr1n.pk0083.c5 csi1.pk0003.g6 p0126.cnlcb49r rls6.pk0068.b7:fis 64 73.2 72.6 72.8

[0111]Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal method of alignment (Higgins and Sharp (1989) CABIOS. 5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5. Sequence alignments and BLAST scores and probabilities indicate that the nucleic acid fragments comprising the instant cDNA clones encode entire corn, rice, and soybean cysteine γ synthases. These sequences represent the first corn, rice, and soybean sequences encoding cysteine γ synthase known to Applicant.

Example 7

Characterization of cDNA Clones Encoding Cystathione β-Lyase

[0112]The BLASTX search using the EST sequences from clones listed in Table 15 revealed similarity of the polypeptides encoded by the cDNAs to cystathionine β-lyase from Arabidopsis thaliana (GenBank Accession No. L40511; NCBI General Identifier No. 1708993). Shown in Table 15 are the BLAST results for individual ESTs ("EST"), the sequences of the entire cDNA inserts comprising the indicated cDNA clones ("FIS"), or the sequences of FISs encoding the entire protein ("CGS"):

TABLE-US-00016 TABLE 15 BLAST Results for Sequences Encoding Polypeptides Homologous to Cystathione β-Lyase BLAST pLog Score Clone Status 1708993 (A. thaliana) cen1.pk0061.d4 FIS 50.41 rlr12.pk0026.g1 EST 39.00 sfl1.pk0012.c4 CGS 33.85 wr1.pk0091.g6 EST 52.52

[0113]The sequence of the entire cDNA insert in the clone wr1.pk0091.g6 was determined, RACE PCR was used to obtain the 5' portion of the rice cystathionine β-lyase, and further sequencing and searching of the DuPont proprietary database allowed the identification of other corn and wheat clones encoding cystathionine β-lyase. The BLASTX search using the EST sequences from clones listed in Table 16 revealed similarity of the polypeptides encoded by the cDNAs to cystathionine β-lyase from Arabidopsis thaliana (GenBank Accession No. L40511; NCBI General Identifier No. 1708993). Shown in Table 16 are the BLAST results for the sequences of the entire cDNA inserts comprising the indicated cDNA clones ("FIS"), or the sequences encoding the entire protein derived from contigs assembled from the sequences of more than two ESTs, the sequence of contigs assembled from the entire cDNA inserts comprising the indicated cDNA clones and 5' RACE PCR or an EST ("Contig*"):

TABLE-US-00017 TABLE 16 BLAST Results for Sequences Encoding Polypeptides Homologous to Cystathione β-Lyase BLAST pLog Score Clone Status 1708993 Contig of: Contig* >180.00 cen1.pk0061.d4 p0005.cbmei71r p0014.ctuui39r p0109.cdadg47r p0125.czaay16r 5' RACE PCR + Contig* 178.00 rlr12.pk0026.g1:fis wr1.pk0091.g6:fis FIS 177.00

[0114]FIG. 6 presents an alignment of the amino acid sequences set forth in SEQ ID NOs:34, 36, 38, 40, 68, 70, and 72 with the Arabidopsis thaliana sequence (NCBI General Identifier No. 1708993; SEQ ID NO:41). The data in Table 17 presents a calculation of the percent identity of the amino acid sequences set forth in SEQ ID NOs:34, 36, 38, 40, 68, 70, and 72 with the Arabidopsis thaliana sequence (NCBI General Identifier No. 1708993; SEQ ID NO:41).

TABLE-US-00018 TABLE 17 Percent Identity of Amino Acid Sequences Deduced From the Nucleotide Sequences of cDNA Clones Encoding Polypeptides Homologous to Cystathione β-Lyase Percent Identity to Clone SEQ ID NO. 1708993 (Arabidopsis thaliana) cen1.pk0061.d4 34 83.0 rlr12.pk0026.g1 36 76.0 sfl1.pk0012.c4 38 72.2 wr1.pk0091.g6 40 71.8 Contig of: 68 66.8 cen1.pk0061.d4 p0005.cbmei71r p0014.ctuui39r p0109.cdadg47r p0125.czaay16r 5' RACE PCR + 70 66.2 rlr12.pk0026.g1:fis wr1.pk0091.g6:fis 72 66.2

[0115]The amino acid sequence set forth in SEQ ID NO:34 is identical to amino acids 248 through 470 of the amino acid sequence set forth in SEQ ID NO:68. The amino acid sequence set forth in SEQ ID NO:36 is identical to amino acids 152 through 226 of the amino acid sequence set forth in SEQ ID NO:70. The amino acid sequence set forth in SEQ ID NO:40 is identical to amino acids 3 through 133 of the amino acid sequence set forth in SEQ ID NO:72.

[0116]Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal method of alignment (Higgins and Sharp (1989) CABIOS. 5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5. Sequence alignments and BLAST scores and probabilities indicate that the nucleic acid fragments comprising the instant cDNA clones encode an entire soybean cystathionine β-lyase, a substantial portion and an entire corn and rice cystathionine β-lyases, a portion and a substantial portion of a wheat cystathionine β-lyase.

Example 8

Expression of Chimeric Genes in Monocot Cells

[0117]A chimeric gene comprising a cDNA encoding the instant polypeptides in sense orientation with respect to the maize 27 kD zein promoter that is located 5' to the cDNA fragment, and the 10 kD zein 3' end that is located 3' to the cDNA fragment, can be constructed. The cDNA fragment of this gene may be generated by polymerase chain reaction (PCR) of the cDNA clone using appropriate oligonucleotide primers. Cloning sites (Nco I or Sma I) can be incorporated into the oligonucleotides to provide proper orientation of the DNA fragment when inserted into the digested vector pML103 as described below. Amplification is then performed in a standard PCR. The amplified DNA is then digested with restriction enzymes Nco I and Sma I and fractionated on an agarose gel. The appropriate band can be isolated from the gel and combined with a 4.9 kb Nco I-Sma I fragment of the plasmid pML103. Plasmid pML103 has been deposited under the terms of the Budapest Treaty at ATCC (American Type Culture Collection, 10801 University Blvd., Manassas, Va. 20110-2209), and bears accession number ATCC 97366. The DNA segment from pML103 contains a 1.05 kb Sal I-Nco I promoter fragment of the maize 27 kD zein gene and a 0.96 kb Sma I-Sal I fragment from the 3' end of the maize 10 kD zein gene in the vector pGem9Zf(+) (Promega). Vector and insert DNA can be ligated at 15° C. overnight, essentially as described (Maniatis). The ligated DNA may then be used to transform E. coli XL1-Blue (Epicurian Coli XL-1 Blue®; Stratagene). Bacterial transformants can be screened by restriction enzyme digestion of plasmid DNA and limited nucleotide sequence analysis using the dideoxy chain termination method (Sequenase® DNA Sequencing Kit; U.S. Biochemical). The resulting plasmid construct would comprise a chimeric gene encoding, in the 5' to 3' direction, the maize 27 kD zein promoter, a cDNA fragment encoding the instant polypeptides, and the 10 kD zein 3' region.

[0118]The chimeric gene described above can then be introduced into corn cells by the following procedure. Immature corn embryos can be dissected from developing caryopses derived from crosses of the inbred corn lines H99 and LH132. The embryos are isolated 10 to 11 days after pollination when they are 1.0 to 1.5 mm long. The embryos are then placed with the axis-side facing down and in contact with agarose-solidified N6 medium (Chu et al. (1975) Sci. Sin. Peking 18:659-668). The embryos are kept in the dark at 27° C. Friable embryogenic callus consisting of undifferentiated masses of cells with somatic proembryoids and embryoids borne on suspensor structures proliferates from the scutellum of these immature embryos. The embryogenic callus isolated from the primary explant can be cultured on N6 medium and sub-cultured on this medium every 2 to 3 weeks.

[0119]The plasmid, p35S/Ac (obtained from Dr. Peter Eckes, Hoechst Ag, Frankfurt, Germany) may be used in transformation experiments in order to provide for a selectable marker. This plasmid contains the Pat gene (see European Patent Publication 0 242 236) which encodes phosphinothricin acetyl transferase (PAT). The enzyme PAT confers resistance to herbicidal glutamine synthetase inhibitors such as phosphinothricin. The pat gene in p35 S/Ac is under the control of the 35S promoter from Cauliflower Mosaic Virus (Odell et al. (1985) Nature 313:810-812) and the 3' region of the nopaline synthase gene from the T-DNA of the Ti plasmid of Agrobacterium tumefaciens.

[0120]The particle bombardment method (Klein et al. (1987) Nature 327:70-73) may be used to transfer genes to the callus culture cells. According to this method, gold particles (1 μm in diameter) are coated with DNA using the following technique. Ten μg of plasmid DNAs are added to 50 μL of a suspension of gold particles (60 mg per mL). Calcium chloride (50 μL of a 2.5 M solution) and spermidine free base (20 μL of a 1.0 M solution) are added to the particles. The suspension is vortexed during the addition of these solutions. After 10 minutes, the tubes are briefly centrifuged (5 sec at 15,000 rpm) and the supernatant removed. The particles are resuspended in 200 μL of absolute ethanol, centrifuged again and the supernatant removed. The ethanol rinse is performed again and the particles resuspended in a final volume of 30 μL of ethanol. An aliquot (5 μL) of the DNA-coated gold particles can be placed in the center of a Kapton® flying disc (Bio-Rad Labs). The particles are then accelerated into the corn tissue with a Biolistic® PDS-1000/He (Bio-Rad Instruments, Hercules Calif.), using a helium pressure of 1000 psi, a gap distance of 0.5 cm and a flying distance of 1.0 cm.

[0121]For bombardment, the embryogenic tissue is placed on filter paper over agarose-solidified N6 medium. The tissue is arranged as a thin lawn and covered a circular area of about 5 cm in diameter. The petri dish containing the tissue can be placed in the chamber of the PDS-1000/He approximately 8 cm from the stopping screen. The air in the chamber is then evacuated to a vacuum of 28 inches of Hg. The macrocarrier is accelerated with a helium shock wave using a rupture membrane that bursts when the He pressure in the shock tube reaches 1000 psi.

[0122]Seven days after bombardment the tissue can be transferred to N6 medium that contains gluphosinate (2 mg per liter) and lacks casein or proline. The tissue continues to grow slowly on this medium. After an additional 2 weeks the tissue can be transferred to fresh N6 medium containing gluphosinate. After 6 weeks, areas of about 1 cm in diameter of actively growing callus can be identified on some of the plates containing the glufosinate-supplemented medium. These calli may continue to grow when sub-cultured on the selective medium.

[0123]Plants can be regenerated from the transgenic callus by first transferring clusters of tissue to N6 medium supplemented with 0.2 mg per liter of 2,4-D. After two weeks the tissue can be transferred to regeneration medium (Fromm et al. (1990) Bio/Technology 8:833-839).

Example 9

Expression of Chimeric Genes in Dicot Cells

[0124]A seed-specific expression cassette composed of the promoter and transcription terminator from the gene encoding the β subunit of the seed storage protein phaseolin from the bean Phaseolus vulgaris (Doyle et al. (1986) J. Biol. Chem. 261:9228-9238) can be used for expression of the instant polypeptides in transformed soybean. The phaseolin cassette includes about 500 nucleotides upstream (5') from the translation initiation codon and about 1650 nucleotides downstream (3') from the translation stop codon of phaseolin. Between the 5' and 3' regions are the unique restriction endonuclease sites Nco I (which includes the ATG translation initiation codon), Sma I, Kpn I and Xba I. The entire cassette is flanked by Hind III sites.

[0125]The cDNA fragment of this gene may be generated by polymerase chain reaction (PCR) of the cDNA clone using appropriate oligonucleotide primers. Cloning sites can be incorporated into the oligonucleotides to provide proper orientation of the DNA fragment when inserted into the expression vector. Amplification is then performed as described above, and the isolated fragment is inserted into a pUC18 vector carrying the seed expression cassette.

[0126]Soybean embryos may then be transformed with the expression vector comprising sequences encoding the instant polypeptides. To induce somatic embryos, cotyledons, 3-5 mm in length dissected from surface sterilized, immature seeds of the soybean cultivar A2872, can be cultured in the light or dark at 26° C. on an appropriate agar medium for 6-10 weeks. Somatic embryos which produce secondary embryos are then excised and placed into a suitable liquid medium. After repeated selection for clusters of somatic embryos which multiplied as early, globular staged embryos, the suspensions are maintained as described below.

[0127]Soybean embryogenic suspension cultures can be maintained in 35 mL liquid media on a rotary shaker, 150 rpm, at 26° C. with florescent lights on a 16:8 hour day/night schedule. Cultures are subcultured every two weeks by inoculating approximately 35 mg of tissue into 35 mL of liquid medium.

[0128]Soybean embryogenic suspension cultures may then be transformed by the method of particle gun bombardment (Klein et al. (1987) Nature (London) 327:70-73, U.S. Pat. No. 4,945,050). A DuPont Biolistic® PDS1000/HE instrument (helium retrofit) can be used for these transformations.

[0129]A selectable marker gene which can be used to facilitate soybean transformation is a chimeric gene composed of the 35S promoter from Cauliflower Mosaic Virus (Odell et al. (1985) Nature 313:810-812), the hygromycin phosphotransferase gene from plasmid pJR225 (from E. coli; Gritz et al.(1983) Gene 25:179-188) and the 3' region of the nopaline synthase gene from the T-DNA of the Ti plasmid of Agrobacterium tumefaciens. The seed expression cassette comprising the phaseolin 5' region, the fragment encoding the instant polypeptides and the phaseolin 3' region can be isolated as a restriction fragment. This fragment can then be inserted into a unique restriction site of the vector carrying the marker gene.

[0130]To 50 μL of a 60 mg/mL 1 μm gold particle suspension is added (in order): 5 μL DNA (1 μg/μL), 20 μL spermidine (0.1 M), and 50 μL CaCl₂ (2.5 M). The particle preparation is then agitated for three minutes, spun in a microfuge for 10 seconds and the supernatant removed. The DNA-coated particles are then washed once in 400 μL 70% ethanol and resuspended in 40 μL of anhydrous ethanol. The DNA/particle suspension can be sonicated three times for one second each. Five μL of the DNA-coated gold particles are then loaded on each macro carrier disk.

[0131]Approximately 300-400 mg of a two-week-old suspension culture is placed in an empty 60×15 mm petri dish and the residual liquid removed from the tissue with a pipette. For each transformation experiment, approximately 5-10 plates of tissue are normally bombarded. Membrane rupture pressure is set at 1100 psi and the chamber is evacuated to a vacuum of 28 inches mercury. The tissue is placed approximately 3.5 inches away from the retaining screen and bombarded three times. Following bombardment, the tissue can be divided in half and placed back into liquid and cultured as described above.

[0132]Five to seven days post bombardment, the liquid media may be exchanged with fresh media, and eleven to twelve days post bombardment with fresh media containing 50 mg/mL hygromycin. This selective media can be refreshed weekly. Seven to eight weeks post bombardment, green, transformed tissue may be observed growing from untransformed, necrotic embryogenic clusters. Isolated green tissue is removed and inoculated into individual flasks to generate new, clonally propagated, transformed embryogenic suspension cultures. Each new line may be treated as an independent transformation event. These suspensions can then be subcultured and maintained as clusters of immature embryos or regenerated into whole plants by maturation and germination of individual somatic embryos.

Example 10

Expression of Chimeric Genes in Microbial Cells

[0133]The cDNAs encoding the instant polypeptides can be inserted into the T7 E. coli expression vector pBT430. This vector is a derivative of pET-3a (Rosenberg et al. (1987) Gene 56:125-135) which employs the bacteriophage T7 RNA polymerase/T7 promoter system. Plasmid pBT430 was constructed by first destroying the EcoR I and Hind III sites in pET-3a at their original positions. An oligonucleotide adaptor containing EcoR I and Hind III sites was inserted at the BamH I site of pET-3a. This created pET-3aM with additional unique cloning sites for insertion of genes into the expression vector. Then, the Nde I site at the position of translation initiation was converted to an Nco I site using oligonucleotide-directed mutagenesis. The DNA sequence of pET-3aM in this region, 5'-CATATGG, was converted to 5'-CCCATGG in pBT430.

[0134]Plasmid DNA containing a cDNA may be appropriately digested to release a nucleic acid fragment encoding the protein. This fragment may then be purified on a 1% low melting agarose gel. Buffer and agarose contain 10 μg/ml ethidium bromide for visualization of the DNA fragment. The fragment can then be purified from the agarose gel by digestion with GELase® (Epicentre Technologies, Madison, Wis.) according to the manufacturer's instructions, ethanol precipitated, dried and resuspended in 20 μL of water. Appropriate oligonucleotide adapters may be ligated to the fragment using T4 DNA ligase (New England Biolabs (NEB), Beverly, Mass.). The fragment containing the ligated adapters can be purified from the excess adapters using low melting agarose as described above. The vector pBT430 is digested, dephosphorylated with alkaline phosphatase (NEB) and deproteinized with phenol/chloroform as described above. The prepared vector pBT430 and fragment can then be ligated at 16° C. for 15 hours followed by transformation into DH5 electrocompetent cells (GIBCO BRL). Transformants can be selected on agar plates containing LB media and 100 μg/mL ampicillin. Transformants containing the gene encoding the instant polypeptides are then screened for the correct orientation with respect to the T7 promoter by restriction enzyme analysis.

[0135]For high level expression, a plasmid clone with the cDNA insert in the correct orientation relative to the T7 promoter can be transformed into E. coli strain BL21(DE3) (Studier et al. (1986) J. Mol. Biol. 189:113-130). Cultures are grown in LB medium containing ampicillin (100 mg/L) at 25° C. At an optical density at 600 nm of approximately 1, IPTG (isopropylthio-β-galactoside, the inducer) can be added to a final concentration of 0.4 mM and incubation can be continued for 3 h at 25°. Cells are then harvested by centrifugation and re-suspended in 50 μL of 50 mM Tris-HCl at pH 8.0 containing 0.1 mM DTT and 0.2 mM phenyl methylsulfonyl fluoride. A small amount of 1 mm glass beads can be added and the mixture sonicated 3 times for about 5 seconds each time with a microprobe sonicator. The mixture is centrifuged and the protein concentration of the supernatant determined. One μg of protein from the soluble fraction of the culture can be separated by SDS-polyacrylamide gel electrophoresis. Gels can be observed for protein bands migrating at the expected molecular weight.

Example 11

Evaluating Compounds for Their Ability to Inhibit the Activity

of Plant Biosynthetic Enzymes

[0136]The polypeptides described herein may be produced using any number of methods known to those skilled in the art. Such methods include, but are not limited to, expression in bacteria as described in Example 10, or expression in eukaryotic cell culture, in planta, and using viral expression systems in suitably infected organisms or cell lines. The instant polypeptides may be expressed either as mature forms of the proteins as observed in vivo or as fusion proteins by covalent attachment to a variety of enzymes, proteins or affinity tags. Common fusion protein partners include glutathione S-transferase ("GST"), thioredoxin ("Trx"), maltose binding protein, and C- and/or N-terminal hexahistidine polypeptide ("(His)₆"). The fusion proteins may be engineered with a protease recognition site at the fusion point so that fusion partners can be separated by protease digestion to yield intact mature enzyme. Examples of such proteases include thrombin, enterokinase and factor Xa. However, any protease can be used which specifically cleaves the peptide connecting the fusion protein and the enzyme.

[0137]Purification of the instant polypeptides, if desired, may utilize any number of separation technologies familiar to those skilled in the art of protein purification. Examples of such methods include, but are not limited to, homogenization, filtration, centrifugation, heat denaturation, ammonium sulfate precipitation, desalting, pH precipitation, ion exchange chromatography, hydrophobic interaction chromatography and affinity chromatography, wherein the affinity ligand represents a substrate, substrate analog or inhibitor. When the instant polypeptides are expressed as fusion proteins, the purification protocol may include the use of an affinity resin which is specific for the fusion protein tag attached to the expressed enzyme or an affinity resin containing ligands which are specific for the enzyme. For example, the instant polypeptides may be expressed as a fusion protein coupled to the C-terminus of thioredoxin. In addition, a (His)₆ peptide may be engineered into the N-terminus of the fused thioredoxin moiety to afford additional opportunities for affinity purification. Other suitable affinity resins could be synthesized by linking the appropriate ligands to any suitable resin such as Sepharose-4B. In an alternate embodiment, a thioredoxin fusion protein may be eluted using dithiothreitol; however, elution may be accomplished using other reagents which interact to displace the thioredoxin from the resin. These reagents include β-mercaptoethanol or other reduced thiol. The eluted fusion protein may be subjected to further purification by traditional means as stated above, if desired. Proteolytic cleavage of the thioredoxin fusion protein and the enzyme may be accomplished after the fusion protein is purified or while the protein is still bound to the ThioBond® affinity resin or other resin.

[0138]Crude, partially purified or purified enzyme, either alone or as a fusion protein, may be utilized in assays for the evaluation of compounds for their ability to inhibit enzymatic activation of the instant polypeptides disclosed herein. Assays may be conducted under well known experimental conditions which permit optimal enzymatic activity. Examples of assays for many of these enzymes can be found in Methods in Enzymology Vol. V, (Colowick and Kaplan eds.) Academic Press, New York or Methods in Enzymology Vol. XVII, (Tabor and Tabor eds.) Academic Press, New York. Specific examples may be found in the following references, each of which is incorporated herein by reference: aspartic semialdehyde dehydrogenase may be assayed as described in Black et al. (1955) J. Biol. Chem. 213:39-50, or Cremer et al. (1988) J. Gen. Microbiol. 134:3221-3229; diaminopimelate decarboxylase may be assayed as described in Work (1962) in Methods in Enzymology Vol. V, (Colowick and Kaplan eds.) 864-870, Academic Press, New York or Cremer et al. (1988) J. Gen. Microbiol. 134:3221-3229; homoserine kinase may be assayed as described in Aarnes (1976) Plant Sci. Lett. 7:187-194; cysteine synthase may be assayed as described in Thompson et al. (1968) Biochem. Biophys. Res. Commun. 31: 281-286 or Bertagnolli et al. (1977) Plant Physiol. 60:115-121; and cystathionine β-lyase may be assayed as described in Giovanelli et al. (1971) Biochim. Biophys. Acta 227:654-670 or Droux et al. (1995) Arch. Biochem Biophys. 316:585-595.

Sequence CWU 1

721826DNAOryza sativa 1tggtaccgcc acgccaaggt ggtaaggatg gttgtcagca cttaccaagc agcaagtggt 60gctggggctg cggccatgga agaactcaaa cttcaaactc aagaggtctt ggcggggaaa 120gcaccaacat gcaacatttt cagtcagcag tatgctttta atatattttc acataatgca 180ccaattgttg aaaatgggta caatgaggag gagatgaaga tggtgaagga gaccagaaaa 240atctggaatg ataaagatgt gaaggtaact gcaacctgca tacgagttcc tgtgatgcgt 300gcacatgctg aaagtgtgaa tctacagttt gaaaagccac ttgatgagga tactgcaagg 360gaaatcttga gggcagctga aggtgttacc attattgatg accgtgcttc caatcgcttc 420cccacacctc ttgaggtatc ggataaagat gatgtagcag tgggtagaat tcgtcaggat 480ttgtcgcaag atgataacaa agggctggac atatttgttt gtggagatca aatacgtaaa 540ggtgctgcac tcaatgctgt gcagattgct gaaatgctac tcaagtgatt ttcttttctg 600tacctttctc tccttgcccc tctttgctct agtcattgtt tgacggatgt actctggtta 660gtatgagatc aattttgatc atcttttgta atctatattc ctagtgaaat aaatgtaaaa 720cggttttgct ctatcttctg cacaagtgta gaagaaatct gaaattggga aattggagtg 780tggcccttgt tcaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaa 8262195PRTOryza sativa 2Trp Tyr Arg His Ala Lys Val Val Arg Met Val Val Ser Thr Tyr Gln1 5 10 15Ala Ala Ser Gly Ala Gly Ala Ala Ala Met Glu Glu Leu Lys Leu Gln20 25 30Thr Gln Glu Val Leu Ala Gly Lys Ala Pro Thr Cys Asn Ile Phe Ser35 40 45Gln Gln Tyr Ala Phe Asn Ile Phe Ser His Asn Ala Pro Ile Val Glu50 55 60Asn Gly Tyr Asn Glu Glu Glu Met Lys Met Val Lys Glu Thr Arg Lys65 70 75 80Ile Trp Asn Asp Lys Asp Val Lys Val Thr Ala Thr Cys Ile Arg Val85 90 95Pro Val Met Arg Ala His Ala Glu Ser Val Asn Leu Gln Phe Glu Lys100 105 110Pro Leu Asp Glu Asp Thr Ala Arg Glu Ile Leu Arg Ala Ala Glu Gly115 120 125Val Thr Ile Ile Asp Asp Arg Ala Ser Asn Arg Phe Pro Thr Pro Leu130 135 140Glu Val Ser Asp Lys Asp Asp Val Ala Val Gly Arg Ile Arg Gln Asp145 150 155 160Leu Ser Gln Asp Asp Asn Lys Gly Leu Asp Ile Phe Val Cys Gly Asp165 170 175Gln Ile Arg Lys Gly Ala Ala Leu Asn Ala Val Gln Ile Ala Glu Met180 185 190Leu Leu Lys1953875DNATriticum aestivum 3cctcatggct gtcacgccgc tgcatcgcca cgccaaggtg aaaaggatgg ttgtcagcac 60ataccaagca gcaagtggtg ctggtgctgc agccatggaa gaactcaaac ttcagactcg 120agaggtcttg gaaggaaagc caccaacctg taacattttc agtcaacagt atgcttttaa 180tatattttcg cataatgcac ctattgttga aaatggctat aatgaggaag agatgaaaat 240ggtgaaggag accagaaaaa tctggaatga caaggatgta agagtaactg caacttgtat 300acgggttcct acgatgcgcg cgcatgccga aagcgtgaat ctacagtttg aaaagccact 360tgatgaggac actgccagag aaatcttgag ggcagctcct ggtgttacca ttagtgacga 420ccgtgctgcc aaccgcttcc ctacaccact ggaggtatcg gataaagatg acgtatcagt 480tggtaggatt cgccaggact tgtcacaaga tgataacaga gggttggagt tatttgtctg 540tggagaccag atacgtaaag gcgccgcgct gaacgctgtg cagattgctg aaatgctact 600gaagtgaccg cctttttacc attgtctcat gtgccacgtt gctctatcca ttgatggatt 660gatgtactct agtcactttc aacccagttt tggtcgtcgt cttttttgta atctgtcaac 720ctagcagaag aagtgtaaga cgggctttag tcatctgttg cacacaaaag tgcagccaca 780agtttagaaa aggagggttt tcacttgttc ggattttgcc ttaggttgga ctttgttgca 840agttgtcgtt tgtttcttga aagctggtct gctgt 8754201PRTTriticum aestivum 4Leu Met Ala Val Thr Pro Leu His Arg His Ala Lys Val Lys Arg Met1 5 10 15Val Val Ser Thr Tyr Gln Ala Ala Ser Gly Ala Gly Ala Ala Ala Met20 25 30Glu Glu Leu Lys Leu Gln Thr Arg Glu Val Leu Glu Gly Lys Pro Pro35 40 45Thr Cys Asn Ile Phe Ser Gln Gln Tyr Ala Phe Asn Ile Phe Ser His50 55 60Asn Ala Pro Ile Val Glu Asn Gly Tyr Asn Glu Glu Glu Met Lys Met65 70 75 80Val Lys Glu Thr Arg Lys Ile Trp Asn Asp Lys Asp Val Arg Val Thr85 90 95Ala Thr Cys Ile Arg Val Pro Thr Met Arg Ala His Ala Glu Ser Val100 105 110Asn Leu Gln Phe Glu Lys Pro Leu Asp Glu Asp Thr Ala Arg Glu Ile115 120 125Leu Arg Ala Ala Pro Gly Val Thr Ile Ser Asp Asp Arg Ala Ala Asn130 135 140Arg Phe Pro Thr Pro Leu Glu Val Ser Asp Lys Asp Asp Val Ser Val145 150 155 160Gly Arg Ile Arg Gln Asp Leu Ser Gln Asp Asp Asn Arg Gly Leu Glu165 170 175Leu Phe Val Cys Gly Asp Gln Ile Arg Lys Gly Ala Ala Leu Asn Ala180 185 190Val Gln Ile Ala Glu Met Leu Leu Lys195 2005457DNAGlycine maxmisc_feature(211)..(211)n is a, c, g, or t 5gtctgtttta aaatccaaca cttaatctct ctcttcgcag cctaaaatcc caatggcttc 60actctctgtt ttgcgccaca accacctctt ctcgggcccc ctcccggccc gccccaagcc 120cacctcctcc tcctcctcca ggatccgaat gtccctccgc gagaacggcc cctccatcgc 180cgtcgtgggc gtcaccggcg ccgtcggcca ngagttcctc tccgtcctct ccgaccgcga 240cttcccctac cgctccattc atatgctggc ttccaagcgc tccgctggac gccgcatcac 300cttcgaggac agggactacn tcttcaggag ctcacgccgg agagttcgac ggtgtcgaca 360tcgcgctctt cagcgcnggg ggtccatcaa nnaagcattc ggaccatcgn cgtaaatcgn 420gggacggncg tngncaanat anctccggtt ncctttg 457686PRTGlycine max 6Met Ala Ser Leu Ser Val Leu Arg His Asn His Leu Phe Ser Gly Pro1 5 10 15Leu Pro Ala Arg Pro Lys Pro Thr Ser Ser Ser Ser Ser Arg Ile Arg20 25 30Met Ser Leu Arg Glu Asn Gly Pro Ser Ile Ala Val Val Gly Val Thr35 40 45Gly Ala Val Gly Gln Glu Phe Leu Ser Val Leu Ser Asp Arg Asp Phe50 55 60Pro Tyr Arg Ser Ile His Met Leu Ala Ser Lys Arg Ser Ala Gly Arg65 70 75 80Arg Ile Thr Phe Glu Asp857160PRTLegionella pneumophila 7Met Ser Arg His Leu Asn Val Ala Ile Val Gly Ala Thr Gly Ala Val1 5 10 15Gly Glu Thr Phe Leu Thr Val Leu Glu Glu Arg Asn Phe Pro Ile Lys20 25 30Ser Leu Tyr Pro Leu Ala Ser Ser Arg Ser Val Gly Lys Thr Val Thr35 40 45Phe Arg Asp Gln Glu Leu Asp Val Leu Asp Leu Ala Glu Phe Asp Phe50 55 60Ser Lys Val Asp Leu Ala Leu Phe Ser Ala Gly Gly Ala Val Ser Lys65 70 75 80Glu Tyr Ala Pro Lys Ala Val Ala Ala Gly Cys Val Val Val Asp Asn85 90 95Thr Ser Cys Phe Arg Tyr Glu Asp Asp Ile Pro Leu Val Val Pro Gly100 105 110Ser Glu Ser Ser Ser Asn Arg Asp Tyr Thr Lys Arg Gly Ile Ile Ala115 120 125Asn Pro Asn Cys Ser Thr Ile Gln Met Val Val Ala Leu Lys Pro Ile130 135 140Tyr Asp Ala Val Gly Ile Ser Arg Ile Asn Val Ala Thr Tyr Gln Ser145 150 155 16081054DNAZea mays 8atttaacgga aatgggaaga cactcgaaca tcttaaatta gctgctgaga gtggagtatt 60tgtaaatgtg gatagcgaat ttgatttgga gaatattgtc agagctgcaa gagctactgg 120aaagaaagtg cctgttttgc ttcgaataaa tccagatgtg gatccgcagg tacatcctta 180tgttgccacg ggaaataaaa cgtctaaatt tgggatccgc aatgagaaat tgcaatggtt 240tttggactct atcaagtcat acccgaatga aatcaaactc gttggtgttc attgccatct 300gggatctact attacaaagg ttgatatatt cagagatgct gcagttctta tgctgaatta 360tgtcgatgaa attcgagcac aaggttttaa gttggagtac ctgaatatcg gaggtggttt 420gggaatagat taccatcata ccgatgcagt cttacctaca cctatggatc tcatcaacac 480tgtgcgagaa ttagttctct ctcaagatct cactcttatt attgaacccg gaagatcctt 540gattgctaat acttgctgct tcgtcaatag agtaactggt gttaaatcta atggtacaaa 600gaatttcatt gttgttgatg gcagcatggc agaactcatc agacctagtc tgtatggagc 660ataccagcat atcgaactgg tctctccccc cactcctggt gctgaagcag cgaccttcga 720tattgttgga ccagtttgtg agtctgcaga tttccttgga aaagataggg aacttccaac 780acctgatgag ggagctggac tggttgttca tgatgcaggt gcctactgca tgagcatggc 840ttccacctac aacctgaagt tgaggccacc ggaatactgg gtggaagcgg acggttcgat 900cgttaagatc aggcatggag agaagcttga tgactacatg aagttctttg atggtcttcc 960tgcttagatg tttattatct gcgactgcta cggacgatgt tttcttgggg ataattggat 1020tttctttgtc aaaaaaaaaa aaaaaaaaaa aaaa 10549321PRTZea mays 9Phe Asn Gly Asn Gly Lys Thr Leu Glu His Leu Lys Leu Ala Ala Glu1 5 10 15Ser Gly Val Phe Val Asn Val Asp Ser Glu Phe Asp Leu Glu Asn Ile20 25 30Val Arg Ala Ala Arg Ala Thr Gly Lys Lys Val Pro Val Leu Leu Arg35 40 45Ile Asn Pro Asp Val Asp Pro Gln Val His Pro Tyr Val Ala Thr Gly50 55 60Asn Lys Thr Ser Lys Phe Gly Ile Arg Asn Glu Lys Leu Gln Trp Phe65 70 75 80Leu Asp Ser Ile Lys Ser Tyr Pro Asn Glu Ile Lys Leu Val Gly Val85 90 95His Cys His Leu Gly Ser Thr Ile Thr Lys Val Asp Ile Phe Arg Asp100 105 110Ala Ala Val Leu Met Leu Asn Tyr Val Asp Glu Ile Arg Ala Gln Gly115 120 125Phe Lys Leu Glu Tyr Leu Asn Ile Gly Gly Gly Leu Gly Ile Asp Tyr130 135 140His His Thr Asp Ala Val Leu Pro Thr Pro Met Asp Leu Ile Asn Thr145 150 155 160Val Arg Glu Leu Val Leu Ser Gln Asp Leu Thr Leu Ile Ile Glu Pro165 170 175Gly Arg Ser Leu Ile Ala Asn Thr Cys Cys Phe Val Asn Arg Val Thr180 185 190Gly Val Lys Ser Asn Gly Thr Lys Asn Phe Ile Val Val Asp Gly Ser195 200 205Met Ala Glu Leu Ile Arg Pro Ser Leu Tyr Gly Ala Tyr Gln His Ile210 215 220Glu Leu Val Ser Pro Pro Thr Pro Gly Ala Glu Ala Ala Thr Phe Asp225 230 235 240Ile Val Gly Pro Val Cys Glu Ser Ala Asp Phe Leu Gly Lys Asp Arg245 250 255Glu Leu Pro Thr Pro Asp Glu Gly Ala Gly Leu Val Val His Asp Ala260 265 270Gly Ala Tyr Cys Met Ser Met Ala Ser Thr Tyr Asn Leu Lys Leu Arg275 280 285Pro Pro Glu Tyr Trp Val Glu Ala Asp Gly Ser Ile Val Lys Ile Arg290 295 300His Gly Glu Lys Leu Asp Asp Tyr Met Lys Phe Phe Asp Gly Leu Pro305 310 315 320Ala101813DNAZea mays 10cgcttcctgg aaggctggaa cagaaagaac cctaaaccct agcaatggcg gcggcgaacc 60tgctgtcgcg ctcccttctc cccaccccaa acactatccg aacgagccac cccaccccgc 120ggagcccagc cgtcgtctcc ttcccccgcc gccgtgcccg cctgtccgtg tgcgcctccg 180tctccatggc ctccccgtcc ccaccgccac agcccgcggc ggccggcgtg ccgaagcact 240gcttccggcg cggcgccgac ggctacctgt actgcgaggg agtgagggtg gaagacgcga 300tggcggctgc cgagcgcagc cccttctatc tctacagcaa gcttcagatc ctccgcaact 360tcgccgctta ccgcgacgct ctccaggggc tccgctccat cgtcgggtat gccgtgaagg 420ccaacaataa cctccccgtg ctacgcgtcc tgcgtgagct tggctgcggc gccgtcctcg 480tcagcggcaa cgagctccga ctcgccctcc aggcgggatt cgaccccgcc aggtgtatat 540ttaacggaaa tgggaagaca ctcgaagatc ttaaattggc tgctgagagt ggagtatttg 600taaatgtgga tagtgaattt gatttagaga atattgtcag agctgcaaga gctactggaa 660agaaagtgcc tgttttactt agaataaatc cagatgtgga tccacaggta catccatatg 720ttgccacggg aaataaaaca tccaaattcg ggatccgcaa tgagaaattg caatggtttt 780tgaactctat caagtcatac tcgaatgaaa tcaaactcgt tggtgttcat tgccatctgg 840gatctactat tacaaaggtt gatatattca gagatgctgc agtgcttatg gtgaattatg 900tcgatgaaat tcgagcacaa ggttttaagt tggagtacct gaatattgga ggtggtttgg 960gaatagatta ccatcatacc gatgcagtct tacctacacc tatggatctc atcaacactg 1020tacgagaatt agttctctct caagatctta ctcttattat tgaacctgga agatccttga 1080ttgctaatac ttgctgcttc gtcaatagag taactggtgt taaatctaat ggtacaaaga 1140atttcattgt tgttgatggc agcatggcag aactcatcag acctagcctg tatggagcat 1200atcagcatat cgaattggtc tctcccccca ctcctggtgc tgaagtagcg accttcgata 1260ttgttgggcc agtttgtgag tctgcagatt tccttggaaa agatagggaa cttccaacac 1320ctgatgaggg agctggactg gttgttcatg atgcaggtgc ctactgcatg agcatggctt 1380ccacctacaa cctgaagttg aggccgccag agtactgggt tgaagaggat ggttcgattg 1440ttaagatcag gcatgaagag aagctcgatg actacatgaa gttctttgat ggtcttcctg 1500cttagatgtt tatttgtgac tgctaggggc gatgttttct tggagataat tgaatttttc 1560tttgtcaagc tcattttgct ttcttgtggt tgttatggaa tgttactgga tactggatag 1620ttagttcggc ctgtaggcgt atcctcctga acttacctct cattgctgtt agttttggca 1680ccaagtttgt tcccaattgc tatttacgga agttattgca taaagggctg tttggttgta 1740atcttcccgt aagaataaga tgcatgtttt tgagttaaaa aagggggggc ccggtaccca 1800attcgcccta tag 181311486PRTZea mays 11Met Ala Ala Ala Asn Leu Leu Ser Arg Ser Leu Leu Pro Thr Pro Asn1 5 10 15Thr Ile Arg Thr Ser His Pro Thr Pro Arg Ser Pro Ala Val Val Ser20 25 30Phe Pro Arg Arg Arg Ala Arg Leu Ser Val Cys Ala Ser Val Ser Met35 40 45Ala Ser Pro Ser Pro Pro Pro Gln Pro Ala Ala Ala Gly Val Pro Lys50 55 60His Cys Phe Arg Arg Gly Ala Asp Gly Tyr Leu Tyr Cys Glu Gly Val65 70 75 80Arg Val Glu Asp Ala Met Ala Ala Ala Glu Arg Ser Pro Phe Tyr Leu85 90 95Tyr Ser Lys Leu Gln Ile Leu Arg Asn Phe Ala Ala Tyr Arg Asp Ala100 105 110Leu Gln Gly Leu Arg Ser Ile Val Gly Tyr Ala Val Lys Ala Asn Asn115 120 125Asn Leu Pro Val Leu Arg Val Leu Arg Glu Leu Gly Cys Gly Ala Val130 135 140Leu Val Ser Gly Asn Glu Leu Arg Leu Ala Leu Gln Ala Gly Phe Asp145 150 155 160Pro Ala Arg Cys Ile Phe Asn Gly Asn Gly Lys Thr Leu Glu Asp Leu165 170 175Lys Leu Ala Ala Glu Ser Gly Val Phe Val Asn Val Asp Ser Glu Phe180 185 190Asp Leu Glu Asn Ile Val Arg Ala Ala Arg Ala Thr Gly Lys Lys Val195 200 205Pro Val Leu Leu Arg Ile Asn Pro Asp Val Asp Pro Gln Val His Pro210 215 220Tyr Val Ala Thr Gly Asn Lys Thr Ser Lys Phe Gly Ile Arg Asn Glu225 230 235 240Lys Leu Gln Trp Phe Leu Asn Ser Ile Lys Ser Tyr Ser Asn Glu Ile245 250 255Lys Leu Val Gly Val His Cys His Leu Gly Ser Thr Ile Thr Lys Val260 265 270Asp Ile Phe Arg Asp Ala Ala Val Leu Met Val Asn Tyr Val Asp Glu275 280 285Ile Arg Ala Gln Gly Phe Lys Leu Glu Tyr Leu Asn Ile Gly Gly Gly290 295 300Leu Gly Ile Asp Tyr His His Thr Asp Ala Val Leu Pro Thr Pro Met305 310 315 320Asp Leu Ile Asn Thr Val Arg Glu Leu Val Leu Ser Gln Asp Leu Thr325 330 335Leu Ile Ile Glu Pro Gly Arg Ser Leu Ile Ala Asn Thr Cys Cys Phe340 345 350Val Asn Arg Val Thr Gly Val Lys Ser Asn Gly Thr Lys Asn Phe Ile355 360 365Val Val Asp Gly Ser Met Ala Glu Leu Ile Arg Pro Ser Leu Tyr Gly370 375 380Ala Tyr Gln His Ile Glu Leu Val Ser Pro Pro Thr Pro Gly Ala Glu385 390 395 400Val Ala Thr Phe Asp Ile Val Gly Pro Val Cys Glu Ser Ala Asp Phe405 410 415Leu Gly Lys Asp Arg Glu Leu Pro Thr Pro Asp Glu Gly Ala Gly Leu420 425 430Val Val His Asp Ala Gly Ala Tyr Cys Met Ser Met Ala Ser Thr Tyr435 440 445Asn Leu Lys Leu Arg Pro Pro Glu Tyr Trp Val Glu Glu Asp Gly Ser450 455 460Ile Val Lys Ile Arg His Glu Glu Lys Leu Asp Asp Tyr Met Lys Phe465 470 475 480Phe Asp Gly Leu Pro Ala485121116DNAOryza sativa 12cttacacgga gtgtttgtaa acatagacag tgaatttgat ttggagaata ttgtcactgc 60tgcgagagtt gctgggaaga aagtccctgt tttgctcagg ataaacccag atgtggatcc 120acaggtccat ccttatgttg cgactggaaa caaaacctcc aaatttggta tccgtaatga 180gaaactacaa tggttcttag actctatcaa gtcatactca aatgatatca cactggtggg 240tgttcattgt catctgggat ctaccattac aaaggtcgat atatttagag atgcggcagg 300tcttatggtg aattatgttg atgaaattcg agcacaaggt tttgaactgg aatatctcaa 360tattggcggt ggcctgggca tagwttatca ccacacggat gcagtcttgc ctacacctat 420gggacctcat caacactgtg ccgaagaatt agttctgtca cgagatctta cactcatcat 480tgaacctggg agatccctca tagctaacac ttgctgcttc gtcaataggg tcactggtgt 540taaatctaat ggtacaaaga atttcattgt agttgatggc agcatggcag agcttatcag 600accaagtcta tatggagcat accagcatat cgaactggtt tctccttccc cagatgcaga 660agtagcaaca ttcgatattg ttggaccagt ttgtgaatct gcagatttcc ttggcaaaga 720cagggaactt ccaacacctg ataagggagc tggtttggtg gttcatgacg caggagccta 780ctgcatgagc atggcttcaa cctacaactt gaagttgcga ccacctgaat attgggtaga 840agatgatggg tccattgcta agattcggcg tggagagtca tttgatgact acatgaagtt 900ctttgataat ctctctgcct aactcgtttt cctgcaattg taataagatt tttctcttgt 960tatgtgtggc tgtatcagga ttcggattga tagcgcagta cagtttgctg tagaatcggt 1020attttttttt attgtactgt gatgtcggta ccttatttta tccaaagatt tttggcaaat 1080tttgctacag gacacttaaa aaaaaaaaaa aaaaaa 111613306PRTOryza sativamisc_feature(128)..(128)Xaa can be any naturally occurring amino acid 13Leu His Gly Val Phe Val Asn Ile Asp Ser Glu Phe Asp Leu Glu Asn1 5 10 15Ile Val Thr Ala Ala Arg Val Ala Gly Lys Lys Val Pro Val Leu Leu20

25 30Arg Ile Asn Pro Asp Val Asp Pro Gln Val His Pro Tyr Val Ala Thr35 40 45Gly Asn Lys Thr Ser Lys Phe Gly Ile Arg Asn Glu Lys Leu Gln Trp50 55 60Phe Leu Asp Ser Ile Lys Ser Tyr Ser Asn Asp Ile Thr Leu Val Gly65 70 75 80Val His Cys His Leu Gly Ser Thr Ile Thr Lys Val Asp Ile Phe Arg85 90 95Asp Ala Ala Gly Leu Met Val Asn Tyr Val Asp Glu Ile Arg Ala Gln100 105 110Gly Phe Glu Leu Glu Tyr Leu Asn Ile Gly Gly Gly Leu Gly Ile Xaa115 120 125Tyr His His Thr Asp Ala Val Leu Pro Thr Pro Met Gly Pro His Gln130 135 140His Cys Ala Glu Glu Leu Val Leu Ser Arg Asp Leu Thr Leu Ile Ile145 150 155 160Glu Pro Gly Arg Ser Leu Ile Ala Asn Thr Cys Cys Phe Val Asn Arg165 170 175Val Thr Gly Val Lys Ser Asn Gly Thr Lys Asn Phe Ile Val Val Asp180 185 190Gly Ser Met Ala Glu Leu Ile Arg Pro Ser Leu Tyr Gly Ala Tyr Gln195 200 205His Ile Glu Leu Val Ser Pro Ser Pro Asp Ala Glu Val Ala Thr Phe210 215 220Asp Ile Val Gly Pro Val Cys Glu Ser Ala Asp Phe Leu Gly Lys Asp225 230 235 240Arg Glu Leu Pro Thr Pro Asp Lys Gly Ala Gly Leu Val Val His Asp245 250 255Ala Gly Ala Tyr Cys Met Ser Met Ala Ser Thr Tyr Asn Leu Lys Leu260 265 270Arg Pro Pro Glu Tyr Trp Val Glu Asp Asp Gly Ser Ile Ala Lys Ile275 280 285Arg Arg Gly Glu Ser Phe Asp Asp Tyr Met Lys Phe Phe Asp Asn Leu290 295 300Ser Ala30514968DNAGlycine max 14gttgccactg ggaataagaa ctctaaattt ggcattagaa atgagaagct gcagtgcttt 60ttagatgcag tgaaggaaca tcctaatgag ctcaaacttg taggggccca ctgccatctt 120ggttcaacaa ttaccaaggt tgacattttc agggatgcag ccaccattat gatcaactac 180attgaccaaa tccgagatca gggttttgaa gttgattact taaatattgg tggaggactt 240gggatagatt attatcattc tggtgccatc cttcctacac ctagagatct cattgacact 300gtacgagatc ttgttatttc acgtggtctt aatctcatca ttgaaccagg aagatcactc 360attgcaaaca cgtgttgctt agttaaccgg gtgacaggtg ttaaaactaa tggatctaaa 420aacttcattg taattgatgg aagtatggct gaacttatcc gccctagtct ttatgatgct 480taccagcata tagagctggt ttcccctgcc ccgtcaaatg ctgaaacaga aacttttgat 540gtggttggcc ctgtctgtga gtctgcagat ttcttaggaa aaggaagaga acttcctact 600ccagccaagg gtactggttt ggttgttcat gatgctggtg cttattgcat gagcatggca 660tcaacctaca atctaaagat gcggcctcct gagtattggg ttgaagatga tggatcagtg 720agcaaaataa gacatggaga gacttttgaa gaccacattc ggttttttga ggggctttga 780gctaataatt tatcttgtag gaaagaaggc tggagaattg ttatgtactt ggagtttgaa 840tctttcctcg tcaatgaatg catgactctt gtagttctgt ttcttccgtt ctaattgaat 900gttgactccc atgacaggaa cagagaataa agttgatttc agttagattt aaaaaaaaaa 960aaaaaaaa 96815259PRTGlycine max 15Val Ala Thr Gly Asn Lys Asn Ser Lys Phe Gly Ile Arg Asn Glu Lys1 5 10 15Leu Gln Cys Phe Leu Asp Ala Val Lys Glu His Pro Asn Glu Leu Lys20 25 30Leu Val Gly Ala His Cys His Leu Gly Ser Thr Ile Thr Lys Val Asp35 40 45Ile Phe Arg Asp Ala Ala Thr Ile Met Ile Asn Tyr Ile Asp Gln Ile50 55 60Arg Asp Gln Gly Phe Glu Val Asp Tyr Leu Asn Ile Gly Gly Gly Leu65 70 75 80Gly Ile Asp Tyr Tyr His Ser Gly Ala Ile Leu Pro Thr Pro Arg Asp85 90 95Leu Ile Asp Thr Val Arg Asp Leu Val Ile Ser Arg Gly Leu Asn Leu100 105 110Ile Ile Glu Pro Gly Arg Ser Leu Ile Ala Asn Thr Cys Cys Leu Val115 120 125Asn Arg Val Thr Gly Val Lys Thr Asn Gly Ser Lys Asn Phe Ile Val130 135 140Ile Asp Gly Ser Met Ala Glu Leu Ile Arg Pro Ser Leu Tyr Asp Ala145 150 155 160Tyr Gln His Ile Glu Leu Val Ser Pro Ala Pro Ser Asn Ala Glu Thr165 170 175Glu Thr Phe Asp Val Val Gly Pro Val Cys Glu Ser Ala Asp Phe Leu180 185 190Gly Lys Gly Arg Glu Leu Pro Thr Pro Ala Lys Gly Thr Gly Leu Val195 200 205Val His Asp Ala Gly Ala Tyr Cys Met Ser Met Ala Ser Thr Tyr Asn210 215 220Leu Lys Met Arg Pro Pro Glu Tyr Trp Val Glu Asp Asp Gly Ser Val225 230 235 240Ser Lys Ile Arg His Gly Glu Thr Phe Glu Asp His Ile Arg Phe Phe245 250 255Glu Gly Leu16676DNATriticum aestivummisc_feature(373)..(373)n is a, c, g, or t 16tttgagttgg agtacctgaa tattggaggt ggtttgggga tagactacca ccacactggt 60gcagtcttgc ctacacctat ggatcttatc aacactgtcc gggaattggt cctctcacgg 120gatcttactc tcattattga acctggaaga tccctgatcg ccaatacttg ctgcttcgtc 180aataaggtca ctggtgtaaa atcgaatggc acgaagaatt tcattgtagt tgatggcagc 240atggccgagc tcatcaggcc tagtctatat ggagcatatc agcatataga actagttctc 300cctctccaag gtgcagaagt agcaaccttc cgatattgtt ggggccagtc tgcgaatctg 360cagattcctt ggnaaagaca aggagttcca acacctgaca aggganctgg tttgggtgtc 420cacgacgcan ganctactgc atgagcatgg cttcnaccta caacctgaag atgaggcaac 480cgagtattgg gtanaggaca tggnccatgt aagataagca cggggaaaca ttgacgacac 540atgagtcttg atngctccgc caggccttta ctggttggna acnagcttca ttgtnnccac 600cgtggaatct gggaacatcn tgttgtagtg gcaccacana gggnttttgn gacaatcaca 660ntagatgaga ttntgg 6761773PRTTriticum aestivum 17Pro Thr Pro Met Asp Leu Ile Asn Thr Val Arg Glu Leu Val Leu Ser1 5 10 15Arg Asp Leu Thr Leu Ile Ile Glu Pro Gly Arg Ser Leu Ile Ala Asn20 25 30Thr Cys Cys Phe Val Asn Lys Val Thr Gly Val Lys Ser Asn Gly Thr35 40 45Lys Asn Phe Ile Val Val Asp Gly Ser Met Ala Glu Leu Ile Arg Pro50 55 60Ser Leu Tyr Gly Ala Tyr Gln His Ile65 7018544DNAGlycine maxmisc_feature(465)..(465)n is a, c, g, or t 18ttgcaacaca cattgtcttg tcggcaaaat cttccaccaa caacacacag ccatggcagg 60ctcaaacatt ctttctcact ctccttccct tcccaaaacc tacagccact ccttaaacca 120aaacgcgtta tcccaaaagc ttttttttct gcccctcaaa ttcaaagcca ccacaaaacc 180acgtgctctc agagcggttc tctcgcagaa cgctgtcaaa acctcggtgg aggacacaaa 240gaacgctcat tttcagcact gtttcaccaa atccgaagat gggtatctgt actgtgaggg 300cctcaaggtg catgacatca tggaatctgt tgagagaaga cctttctatt tgtacagcaa 360gccccagata actaggaatg ttgaagccta caaggatgca ttggaagggt tgaactccat 420aattggttat gccattaagg ccaataataa cttgaagatt ttggnacatt tgaggcactt 480gggttgtggt gctgtgcttg ttagtgggaa tgagctgaag ttgntcttcg agctggnttt 540gttc 5441962PRTGlycine maxmisc_feature(44)..(44)Xaa can be any naturally occurring amino acid 19Arg Arg Pro Phe Tyr Leu Tyr Ser Lys Pro Gln Ile Thr Arg Asn Val1 5 10 15Glu Ala Tyr Lys Asp Ala Leu Glu Gly Leu Asn Ser Ile Ile Gly Tyr20 25 30Ala Ile Lys Ala Asn Asn Asn Leu Lys Ile Leu Xaa His Leu Arg His35 40 45Leu Gly Cys Gly Ala Val Leu Val Ser Gly Asn Glu Leu Lys50 55 6020371PRTPseudomonas aeruginosa 20Met Lys Arg Val Gly Leu Ile Gly Trp Arg Gly Met Val Gly Ser Val1 5 10 15Leu Ile Gln Arg Met Leu Glu Glu Arg Asp Phe Asp Leu Ile Glu Pro20 25 30Val Phe Phe Thr Thr Ser Asn Val Gly Ala Gln Ala Pro Glu Val Asp35 40 45Lys Asp Ile Ala Pro Leu Lys Asp Ala Tyr Ser Ile Asp Glu Leu Lys50 55 60Thr Leu Asp Val Ile Leu Thr Cys Gln Gly Gly Asp Tyr Thr Ser Glu65 70 75 80Val Phe Pro Lys Leu Arg Glu Ala Gly Trp Gln Gly Tyr Trp Ile Asp85 90 95Ala Ala Ser Ser Leu Arg Met Glu Asp Asp Ala Val Ile Val Leu Asp100 105 110Pro Val Asn Arg Lys Val Ile Asp Gln Ala Leu Asp Ala Gly Thr Arg115 120 125Asn Tyr Ile Gly Gly Asn Cys Thr Val Ser Leu Met Leu Met Ala Leu130 135 140Gly Gly Leu Phe Asp Ala Gly Leu Val Glu Trp Met Ser Ala Met Thr145 150 155 160Tyr Gln Ala Ala Ser Gly Ala Gly Ala Gln Asn Met Arg Asp Leu Leu165 170 175Lys Gln Met Gly Ala Ala His Ala Ser Val Ala Asp Asp Leu Ala Asn180 185 190Pro Ala Ser Ala Ile Leu Asp Ile Asp Arg Lys Val Ala Glu Thr Leu195 200 205Arg Ser Glu Ala Phe Pro Thr Glu His Phe Gly Ala Pro Leu Gly Gly210 215 220Ser Leu Ile Pro Trp Ile Asp Lys Glu Leu Ser Gln Arg Arg Gln Ser225 230 235 240Arg Glu Glu Trp Lys Ala Gln Ala Glu Thr Asn Lys Ile Leu Ala Arg245 250 255Phe Lys Asn Pro Ile Pro Val Asp Gly Ile Cys Val Arg Val Gly Ala260 265 270Met Arg Cys His Ser Gln Ala Leu Thr Ile Lys Leu Asn Lys Asp Val275 280 285Pro Leu Thr Asp Ile Glu Gly Leu Ile Arg Gln His Asn Pro Trp Val290 295 300Lys Leu Val Pro Asn His Arg Glu Val Ser Val Arg Glu Leu Thr Pro305 310 315 320Ala Ala Val Thr Gly Thr Leu Ser Val Pro Val Gly Arg Leu Arg Lys325 330 335Leu Asn Met Val Ser Gln Tyr Leu Gly Ala Phe Thr Val Gly Asp Gln340 345 350Leu Leu Trp Gly Ala Ala Glu Pro Leu Arg Arg Met Leu Arg Ile Leu355 360 365Leu Glu Arg37021788DNAZea mays 21cgacaacatc gcccccgcca tcctcggcgg cttcgtcctc gtccgcagct acgacccctt 60tcacctcgtc ccgctttcct tcccgccagc gctccgcctc cacttcgtcc tggtcacccc 120cgacttcgag gcgcccacga gcaagatgcg cgccgcgctg cccaggcagg tcgacgtcca 180gcagcacgtg cgcaactcca gccaggcagc ggcgctcgtg gcggcggtgc tgcaggggga 240cgcgggcctc atcggctccg cgatgtcgtc cgacggcatc gtggagccca ccagggcacc 300cctcatacct ggcatggcgg ccgtaaaggc ggcggccctg caagctggag cgctgggctg 360cacaattagc ggcgcgggcc ccacagtggt ggccgtcatc caaggggagg aaagggggga 420ggaggttgcc cgcaagatgg tggacgcgtt ctggagcgca ggcaagctca aggcgacagc 480aaccgtcgcg cagctcgata cccttggtgc cagggtcatc gccacgtcat ccttgaacta 540gcaaaagatt cggaaagtgg tactgcaatt gtatcaccaa acaaggaaga atgaagggga 600accccatgga tttgtatgtt ttctcttctt tcttgcatct ttaggtggtt aattggcttt 660ggaataaatg agatggagga catcgctaga acaattctgt tccgtgggct gtaatttcaa 720tttgggctgg tttctttatc atgccatgga taattatgaa taaatttgag gtagtttgtt 780aaaaaaaa 78822179PRTZea mays 22Asp Asn Ile Ala Pro Ala Ile Leu Gly Gly Phe Val Leu Val Arg Ser1 5 10 15Tyr Asp Pro Phe His Leu Val Pro Leu Ser Phe Pro Pro Ala Leu Arg20 25 30Leu His Phe Val Leu Val Thr Pro Asp Phe Glu Ala Pro Thr Ser Lys35 40 45Met Arg Ala Ala Leu Pro Arg Gln Val Asp Val Gln Gln His Val Arg50 55 60Asn Ser Ser Gln Ala Ala Ala Leu Val Ala Ala Val Leu Gln Gly Asp65 70 75 80Ala Gly Leu Ile Gly Ser Ala Met Ser Ser Asp Gly Ile Val Glu Pro85 90 95Thr Arg Ala Pro Leu Ile Pro Gly Met Ala Ala Val Lys Ala Ala Ala100 105 110Leu Gln Ala Gly Ala Leu Gly Cys Thr Ile Ser Gly Ala Gly Pro Thr115 120 125Val Val Ala Val Ile Gln Gly Glu Glu Arg Gly Glu Glu Val Ala Arg130 135 140Lys Met Val Asp Ala Phe Trp Ser Ala Gly Lys Leu Lys Ala Thr Ala145 150 155 160Thr Val Ala Gln Leu Asp Thr Leu Gly Ala Arg Val Ile Ala Thr Ser165 170 175Ser Leu Asn23601DNAOryza sativamisc_feature(433)..(433)n is a, c, g, or t 23gtcgccgcca tcgctgccct tcgcgccctc gatgtcaagt cccacgccgt ctccatccac 60ctcaccaagg gcctccccct cggctccggc ctcggctcct ccgccgcctc cgccgccgcc 120gctgccaagg ccgttgacgc cctcttcggc tccctcctac accaagatga cctcgtcctc 180gcgggcctcg agtccgagaa agccgtcagt ggcttccacg ccgacaacat cgccccggcc 240atcctcggcg gcttcgtcct cgtccgcagc tacgacccct tccacctcat cccgctctcc 300tccccacctg ccctccgcct ccacttcgtc ctcgtcacgc ccgacttcga ggcgcccacc 360aagcaagatg cgtgccgcgc tgcccaaaca ggtggccgtc caccaagcac gtccgcaact 420ccagccaagc ggncgcgctt gtcgccgctg tgctgcaagg ggacgccacc ctcatcggct 480ccgcaatgtc ctccgacggc atcgtggagc caacaaggcg ccgctgattc tggatggctg 540cggtcaaagg cgccggcttg gaactggggg aattggctgc acatcagtgg agaaggcaan 600t 6012482PRTOryza sativamisc_feature(56)..(57)Xaa can be any naturally occurring amino acid 24Val Ser Ile His Leu Thr Lys Gly Leu Pro Leu Gly Ser Gly Leu Gly1 5 10 15Ser Ser Ala Ala Ser Ala Ala Ala Ala Ala Lys Ala Val Asp Ala Leu20 25 30Phe Gly Ser Leu Leu His Gln Asp Asp Leu Val Leu Ala Gly Leu Glu35 40 45Ser Glu Lys Ala Val Ser Gly Xaa Xaa His Ala Asp Asn Ile Ala Pro50 55 60Ala Ile Leu Gly Gly Phe Val Leu Val Arg Ser Tyr Asp Pro Phe His65 70 75 80Leu Ile251543DNAGlycine max 25gaagagagac aaaccagcaa gagtggagat ggcgacgtcg acgtgcttcc tgtgtccgtc 60tacggcgagt ttgaaaggca gggccagatt cagaatcaga atcagatgca gcagcagcgt 120gtcggtcaat attcgaaggg agcccgaacc tgtaacgacg ctggtgaaag cgtttgctcc 180cgccacggtg gcgaatctag gtccaggctt cgacttccta ggctgcgccg tggacggact 240cggagacatt gtgtcggtga aggttgaccc acaggttcac cctggcgaga tatgcatatc 300cgacatcagc ggccacgccc caaacaagct cagcaaaaac cctctctgga actgcgccgg 360catcgccgcc attgaagtca tgaaaatgct ctccattcga tccgtcggcc tctccctctc 420cctggagaag ggcctgcctt tgggaagcgg tctgggatcc agcgccgcca gcgccgccgc 480ggccgccgtg gcggtgaacg agctgtttgg gaagaaatta agcgtggagg agctggttct 540ggcatcactg aaatcggaag agaaggtgtc ggggtatcac gcggacaacg tggcgccatc 600gataatgggg ggttttgtgc tgatcgggag ctactcgccg ctggagttga tgccgttgaa 660gtttccggca gagaaggagc tgtatttcgt gctggtgacg cctgagttcg aggccccgac 720gaagaagatg cgggcagcgc tgcctacgga gatcgggatg ccgcaccacg tgtggaactg 780cagccaggca ggtgctctgg tggcgtcggt gctgcagggc gacgtggttg ggttggggaa 840ggcattgtcc tctgacaaga tcgttgagcc aaggcgtgcc cccttgattc ctggcatgga 900ggctgtcaag agggctgcca ttcaggccgg tgcttttggc tgtaccatca gcggcgccgg 960ccctaccgcc gtcgccgtca ttgacgacga gcaaactgga cacctcattg ccaaacacat 1020gattgacgct tttctccatg ttggcaattt gaaggcttct gcaaatgtca agcagcttga 1080tcgccttggt gctagacgca ttccaaattg aaccttctct tctctatctc tatgagaggc 1140ttgtagattt caagaaccgg atttcttcca acttgctcgt aacactctaa gtgctgaccg 1200gtcacatgta tttgaaattt gatctgatca atgaagcagc attctagtgt ggaggtctga 1260ataacaagag aaacattaaa cccaagctgg gagctctgtt tgggtggtgg aaatttaaat 1320agatgaataa ttatgaaaga cctagatcag gtcagtgtta tggtgaactc tgaagcatgt 1380tttagatttt ctttgctttg tttttatcat atttttatct tgctacttga gttgacaaag 1440ctcaaaaaga agtcattttt agtattttct tgtttcatta tgctagttaa tcttagcttt 1500tgaatagcat gtattgttcc ttaaaaaaaa aaaaaaaaaa aaa 154326483PRTGlycine max 26Met Ala Thr Ser Thr Cys Phe Leu Cys Pro Ser Thr Ala Ser Leu Lys1 5 10 15Gly Arg Ala Arg Phe Arg Ile Arg Ile Arg Cys Ser Ser Ser Val Ser20 25 30Val Asn Ile Arg Arg Glu Pro Glu Pro Val Thr Thr Leu Val Lys Ala35 40 45Phe Ala Pro Ala Thr Val Ala Asn Leu Gly Pro Gly Phe Asp Phe Leu50 55 60Gly Cys Ala Val Asp Gly Leu Gly Asp Ile Val Ser Val Lys Val Asp65 70 75 80Pro Gln Val His Pro Gly Glu Ile Cys Ile Ser Asp Ile Ser Gly His85 90 95Ala Pro Asn Lys Leu Ser Lys Asn Pro Leu Trp Asn Cys Ala Gly Ile100 105 110Ala Ala Ile Glu Val Met Lys Met Leu Ser Ile Arg Ser Val Gly Leu115 120 125Ser Leu Ser Leu Glu Lys Gly Leu Pro Leu Gly Ser Gly Leu Gly Ser130 135 140Ser Ala Ala Ser Ala Ala Ala Ala Ala Val Ala Val Asn Glu Leu Phe145 150 155 160Gly Lys Lys Leu Ser Val Glu Glu Leu Val Leu Ala Ser Leu Lys Ser165 170 175Glu Glu Lys Val Ser Gly Tyr His Ala Asp Asn Val Ala Pro Ser Ile180 185 190Met Gly Gly Phe Val Leu Ile Gly Ser Tyr Ser Pro Leu Glu Leu Met195 200 205Pro Leu Lys Phe Pro Ala Glu Lys Glu Leu Tyr Phe Val Leu Val Thr210 215 220Pro Glu Phe Glu Ala Pro Thr Lys Lys Met Arg Ala Ala Leu Pro Thr225 230 235 240Glu Ile Gly Met Pro His His Val Trp Asn Cys Ser Gln Ala Gly Ala245 250 255Leu Val Ala Ser Val Leu Gln Gly Asp Val Val Gly Leu Gly Lys Ala260 265 270Leu Ser Ser Asp Lys Ile Val Glu Pro Arg Arg Ala Pro Leu Ile Pro275 280 285Gly Met Glu Ala Val Lys Arg Ala Ala Ile Gln Ala Gly Ala Phe Gly290 295 300Cys Thr Ile Ser Gly Ala Gly Pro Thr Ala Val Ala Val Ile Asp Asp305 310 315

320Glu Gln Thr Gly His Leu Ile Ala Lys His Met Ile Asp Ala Phe Leu325 330 335His Val Gly Asn Leu Lys Ala Ser Ala Asn Val Lys Gln Leu Asp Arg340 345 350Leu Gly Ala Arg Arg Ile Pro Asn Thr Phe Ser Ser Leu Ser Leu Glu355 360 365Ala Cys Arg Phe Gln Glu Pro Asp Phe Phe Gln Leu Ala Arg Asn Thr370 375 380Leu Ser Ala Asp Arg Ser His Val Phe Glu Ile Ser Asp Gln Ser Ser385 390 395 400Ile Leu Val Trp Arg Ser Glu Gln Glu Lys His Thr Gln Ala Gly Ser405 410 415Ser Val Trp Val Val Glu Ile Ile Asp Glu Leu Lys Thr Ile Arg Ser420 425 430Val Leu Trp Thr Leu Lys His Val Leu Asp Phe Leu Cys Phe Val Phe435 440 445Ile Ile Phe Leu Ser Cys Tyr Leu Ser Gln Ser Ser Lys Arg Ser His450 455 460Phe Tyr Phe Leu Val Ser Leu Cys Leu Ile Leu Ala Phe Glu His Val465 470 475 480Leu Phe Leu27438DNATriticum aestivummisc_feature(271)..(271)n is a, c, g, or t 27ctcgagtcgg agaaggccgt cagcggcttc cacgccgaca acatcgcccc cgccatcctc 60ggcggcttcg tcctcgtccg cagctacgac ccctttcacc tcgtcccgct ttccttcccg 120ccagcgctcc gcctccactt cgtcctggtc acccccgact tcgaggcgcc cacgagcaag 180atgcgcgccg cgctgcccag gcaggtcgac gtccagcagc acgtgcgcaa ctccagccag 240gcagcggcgc tccgtggcgg cggtgctgca nggggacgcc gggctcatcg gtccgcgatt 300tctccgacgg gcatcgtgga cccaccaagg aaccctcata cctggcatgg cggccgtaaa 360ggcggcggcc tgcaactgga cgctgggtgc acattaacgg gcgggcccac atggtggctc 420ncagngaaga gaggggag 4382884PRTTriticum aestivum 28Leu Glu Ser Glu Lys Ala Val Ser Gly Phe His Ala Asp Asn Ile Ala1 5 10 15Pro Ala Ile Leu Gly Gly Phe Val Leu Val Arg Ser Tyr Asp Pro Phe20 25 30His Leu Val Pro Leu Ser Phe Pro Pro Ala Leu Arg Leu His Phe Val35 40 45Leu Val Thr Pro Asp Phe Glu Ala Pro Thr Ser Lys Met Arg Ala Ala50 55 60Leu Pro Arg Gln Val Asp Val Gln Gln His Val Arg Asn Ser Ser Gln65 70 75 80Ala Ala Ala Leu29300PRTMethanococcus jannashii 29Met Arg Glu Ile Met Lys Val Arg Val Lys Ala Pro Cys Thr Ser Ala1 5 10 15Asn Leu Gly Val Gly Phe Asp Val Phe Gly Leu Cys Leu Lys Glu Pro20 25 30Tyr Asp Val Ile Glu Val Glu Ala Ile Asp Asp Lys Glu Ile Ile Ile35 40 45Glu Val Asp Asp Lys Asn Ile Pro Thr Asp Pro Asp Lys Asn Val Ala50 55 60Gly Ile Val Ala Lys Lys Met Ile Asp Asp Phe Asn Ile Gly Lys Gly65 70 75 80Val Lys Ile Thr Ile Lys Lys Gly Val Lys Ala Gly Ser Gly Leu Gly85 90 95Ser Ser Ala Ala Ser Ser Ala Gly Thr Ala Tyr Ala Ile Asn Glu Leu100 105 110Phe Lys Leu Asn Leu Asp Lys Leu Lys Leu Val Asp Tyr Ala Ser Tyr115 120 125Gly Glu Leu Ala Ser Ser Gly Ala Lys His Ala Asp Asn Val Ala Pro130 135 140Ala Ile Phe Gly Gly Phe Thr Met Val Thr Asn Tyr Glu Pro Leu Glu145 150 155 160Val Leu His Ile Pro Ile Asp Phe Lys Leu Asp Ile Leu Ile Ala Ile165 170 175Pro Asn Ile Ser Ile Asn Thr Lys Glu Ala Arg Glu Ile Leu Pro Lys180 185 190Ala Val Gly Leu Lys Asp Leu Val Asn Asn Val Gly Lys Ala Cys Gly195 200 205Met Val Tyr Ala Leu Tyr Asn Lys Asp Lys Ser Leu Phe Gly Arg Tyr210 215 220Met Met Ser Asp Lys Val Ile Glu Pro Val Arg Gly Lys Leu Ile Pro225 230 235 240Asn Tyr Phe Lys Ile Lys Glu Glu Val Lys Asp Lys Val Tyr Gly Ile245 250 255Thr Ile Ser Gly Ser Gly Pro Ser Ile Ile Ala Phe Pro Lys Glu Glu260 265 270Phe Ile Asp Glu Val Glu Asn Ile Leu Arg Asp Tyr Tyr Glu Asn Thr275 280 285Ile Arg Thr Glu Val Gly Lys Gly Val Glu Val Val290 295 300301362DNAGlycine max 30actttgtagt tcgtagatag ccgatgtgct tgtcttagtg tgtcagtcat tcctgttcct 60caagtcaagc tttgtagtga gcagatataa tggctgttga aaggtccgga attgccaaag 120atgttacgga attgattggt aaaaccccat tagtatatct aaataaactt gcggatggtt 180gtgttgcccg ggttgctgct aaactggagt tgatggagcc atgctctagt gtgaaggaca 240ggattgggta tagtatgatt gctgatgcag aagagaaggg acttatcaca cctggaaaga 300gtgtcctcat tgagccaaca agtggtaata ctggcattgg attagccttc atggcagcag 360ccaggggtta caagctcata attacaatgc ctgcttctat gagtcttgag agaagaatca 420ttctattagc ttttggagct gagttggttc tgacagatcc tgctaaggga atgaaaggtg 480ctgttcagaa ggctgaagag atattggcta agacgcccaa tgcctacata cttcaacaat 540ttgaaaaccc tgccaatccc aaggttcatt atgaaaccac tggtccagag atatggaaag 600gctccgatgg gaaaattgat gcatttgttt ctgggatagg cactggtggt acaataacag 660gtgctggaaa atatcttaaa gagcagaatc cgaatataaa gctgattggt gtggaaccag 720ttgaaagtcc agtgctctca ggaggaaagc ctggtccaca caagattcaa gggattggtg 780ctggttttat ccctggtgtc ttggaagtca atcttcttga tgaagttgtt caaatatcaa 840gtgatgaagc aatagaaact gcaaagcttc ttgcgcttaa agaaggccta tttgtgggaa 900tatcttccgg agctgcagct gctgctgctt ttcagattgc aaaaagacca gaaaatgccg 960ggaagcttat tgttgccgtt tttcccagct tcggggagag gtacctgtcc tccgtgctat 1020ttgagtcagt gagacgcgaa gctgaaagca tgacttttga gccctgaatt cccgtttaag 1080gctctcacta ctgaattttc ttgttacttg taccaggctt taactagatt gttagagtac 1140tactgtttgt gactctgact ctaaaataaa acttgctcca aaagactagt ttttcttgat 1200gcccctggag cgataatttt gtgcctgcaa cattaaaaag tattcaaagt tgcttataag 1260taacatgttt catcttttgt tgttgttgag acgaacacgg atgaggtcat aatactatgt 1320ttctgatttc ctttggtagg gaaaaaaaaa aaaaaaaaaa aa 136231325PRTGlycine max 31Met Ala Val Glu Arg Ser Gly Ile Ala Lys Asp Val Thr Glu Leu Ile1 5 10 15Gly Lys Thr Pro Leu Val Tyr Leu Asn Lys Leu Ala Asp Gly Cys Val20 25 30Ala Arg Val Ala Ala Lys Leu Glu Leu Met Glu Pro Cys Ser Ser Val35 40 45Lys Asp Arg Ile Gly Tyr Ser Met Ile Ala Asp Ala Glu Glu Lys Gly50 55 60Leu Ile Thr Pro Gly Lys Ser Val Leu Ile Glu Pro Thr Ser Gly Asn65 70 75 80Thr Gly Ile Gly Leu Ala Phe Met Ala Ala Ala Arg Gly Tyr Lys Leu85 90 95Ile Ile Thr Met Pro Ala Ser Met Ser Leu Glu Arg Arg Ile Ile Leu100 105 110Leu Ala Phe Gly Ala Glu Leu Val Leu Thr Asp Pro Ala Lys Gly Met115 120 125Lys Gly Ala Val Gln Lys Ala Glu Glu Ile Leu Ala Lys Thr Pro Asn130 135 140Ala Tyr Ile Leu Gln Gln Phe Glu Asn Pro Ala Asn Pro Lys Val His145 150 155 160Tyr Glu Thr Thr Gly Pro Glu Ile Trp Lys Gly Ser Asp Gly Lys Ile165 170 175Asp Ala Phe Val Ser Gly Ile Gly Thr Gly Gly Thr Ile Thr Gly Ala180 185 190Gly Lys Tyr Leu Lys Glu Gln Asn Pro Asn Ile Lys Leu Ile Gly Val195 200 205Glu Pro Val Glu Ser Pro Val Leu Ser Gly Gly Lys Pro Gly Pro His210 215 220Lys Ile Gln Gly Ile Gly Ala Gly Phe Ile Pro Gly Val Leu Glu Val225 230 235 240Asn Leu Leu Asp Glu Val Val Gln Ile Ser Ser Asp Glu Ala Ile Glu245 250 255Thr Ala Lys Leu Leu Ala Leu Lys Glu Gly Leu Phe Val Gly Ile Ser260 265 270Ser Gly Ala Ala Ala Ala Ala Ala Phe Gln Ile Ala Lys Arg Pro Glu275 280 285Asn Ala Gly Lys Leu Ile Val Ala Val Phe Pro Ser Phe Gly Glu Arg290 295 300Tyr Leu Ser Ser Val Leu Phe Glu Ser Val Arg Arg Glu Ala Glu Ser305 310 315 320Met Thr Phe Glu Pro32532325PRTCitrullus lanatus 32Met Ala Asp Ala Lys Ser Thr Ile Ala Lys Asp Val Thr Glu Leu Ile1 5 10 15Gly Asn Thr Pro Leu Val Tyr Leu Asn Arg Val Val Asp Gly Cys Val20 25 30Ala Arg Val Ala Ala Lys Leu Glu Met Met Glu Pro Cys Ser Ser Val35 40 45Lys Asp Arg Ile Gly Tyr Ser Met Ile Ser Asp Ala Glu Asn Lys Gly50 55 60Leu Ile Thr Pro Gly Glu Ser Val Leu Ile Glu Pro Thr Ser Gly Asn65 70 75 80Thr Gly Ile Gly Leu Ala Phe Ile Ala Ala Ala Lys Gly Tyr Arg Leu85 90 95Ile Ile Cys Met Pro Ala Ser Met Ser Leu Glu Arg Arg Thr Ile Leu100 105 110Arg Ala Phe Gly Ala Glu Leu Val Leu Thr Asp Pro Ala Arg Gly Met115 120 125Lys Gly Ala Val Gln Lys Ala Glu Glu Ile Lys Ala Lys Thr Pro Asn130 135 140Ser Tyr Ile Leu Gln Gln Phe Glu Asn Pro Ala Asn Pro Lys Ile His145 150 155 160Tyr Glu Thr Thr Gly Pro Glu Ile Trp Arg Gly Ser Gly Gly Lys Ile165 170 175Asp Ala Leu Val Ser Gly Ile Gly Thr Gly Gly Thr Val Thr Gly Ala180 185 190Gly Lys Tyr Leu Lys Glu Gln Asn Pro Asn Ile Lys Leu Tyr Gly Val195 200 205Glu Pro Val Glu Ser Ala Ile Leu Ser Gly Gly Lys Pro Gly Pro His210 215 220Lys Ile Gln Gly Ile Gly Ala Gly Phe Ile Pro Gly Val Leu Asp Val225 230 235 240Asn Leu Leu Asp Glu Val Ile Gln Val Ser Ser Glu Glu Ser Ile Glu245 250 255Thr Ala Lys Leu Leu Ala Leu Lys Glu Gly Leu Leu Val Gly Ile Ser260 265 270Ser Gly Ala Ala Ala Ala Ala Ala Ile Arg Ile Ala Lys Arg Pro Glu275 280 285Asn Ala Gly Lys Leu Ile Val Ala Val Phe Pro Ser Phe Gly Glu Arg290 295 300Tyr Leu Ser Thr Val Leu Phe Glu Ser Val Lys Arg Glu Thr Glu Asn305 310 315 320Met Val Phe Glu Pro32533789DNAZea mays 33atagcgcatt ctcatggtgc tcttgttttg gttgacaaca gcatcatgtc tccagtgctc 60tcccgtccta tagaactggg agctgatatc gtgatgcact cggctaccaa atttatagcg 120ggacatagtg atcttatggc tggaattctt gcagtgaagg gtgagagttt ggctaaagag 180gtagggtttc tgcaaaatgc tgaagggtcg ggtctggcac cttttgactg ctggctttgc 240ttgaggggaa tcaaaaccat ggctctgcgg gtggagaaac aacaggctaa tgcccagaag 300attgctgaat tcctggcgtc tcacccgagg gtcaagcaag taaactacgc tgggcttcct 360gaccatcctg ggcgagcttt acactattcc caggcaaagg gagcgggctc tgttctcagt 420tttctcaccg gctcactggc cctctcaaag cacgtcgtgg agaccaccaa gtacttcagc 480gtaacagtca gcttcgggag cgtgaagtcc ctcatcagcc tgccgtgctt catgtcccac 540gcatcaatcc ctgcctcggt ccgcgaggag cgtggcctaa ccgacgacct cgtccggata 600tcggtcggca tcgaggatgt cgaggacctc atcgccgatc tggaccgcgc gctcagaact 660ggcccggtgt agacatcgcc gatccttagg tcatgtcaag ctatcttttg atgattcatt 720ggttgactgc ttgcgtgatg ataataatgg gaatgttgct tggataaaaa aaaaaaaaaa 780aaaactcga 78934223PRTZea mays 34Ile Ala His Ser His Gly Ala Leu Val Leu Val Asp Asn Ser Ile Met1 5 10 15Ser Pro Val Leu Ser Arg Pro Ile Glu Leu Gly Ala Asp Ile Val Met20 25 30His Ser Ala Thr Lys Phe Ile Ala Gly His Ser Asp Leu Met Ala Gly35 40 45Ile Leu Ala Val Lys Gly Glu Ser Leu Ala Lys Glu Val Gly Phe Leu50 55 60Gln Asn Ala Glu Gly Ser Gly Leu Ala Pro Phe Asp Cys Trp Leu Cys65 70 75 80Leu Arg Gly Ile Lys Thr Met Ala Leu Arg Val Glu Lys Gln Gln Ala85 90 95Asn Ala Gln Lys Ile Ala Glu Phe Leu Ala Ser His Pro Arg Val Lys100 105 110Gln Val Asn Tyr Ala Gly Leu Pro Asp His Pro Gly Arg Ala Leu His115 120 125Tyr Ser Gln Ala Lys Gly Ala Gly Ser Val Leu Ser Phe Leu Thr Gly130 135 140Ser Leu Ala Leu Ser Lys His Val Val Glu Thr Thr Lys Tyr Phe Ser145 150 155 160Val Thr Val Ser Phe Gly Ser Val Lys Ser Leu Ile Ser Leu Pro Cys165 170 175Phe Met Ser His Ala Ser Ile Pro Ala Ser Val Arg Glu Glu Arg Gly180 185 190Leu Thr Asp Asp Leu Val Arg Ile Ser Val Gly Ile Glu Asp Val Glu195 200 205Asp Leu Ile Ala Asp Leu Asp Arg Ala Leu Arg Thr Gly Pro Val210 215 22035547DNAOryza sativamisc_feature(260)..(260)n is a, c, g, or t 35gccttatggc taagcttgag aaggcggatc aggcattctg cttcaccagt gggatggcag 60cactagctgc agtaacacac ctccttaagt ctggacaaga aatagttgct ggagaggaca 120tatatggtgg ctcagaccgt ctgctctcac aagttgcccc gagacatggg attgtagtaa 180aacgaattga tacaaccaaa attagtgagg taacttctgc aattggggcc ttggactaaa 240ctaagtatgg ctttgaaaan cccaccatcc ccgtcctaca aattactgga tataaagaaa 300atagcnagag atagtcatta caatggggct ccttgtttta agtagacaac agcacatgtc 360tccctgtgct ctcccngtcc tcntaaaact ttgggccaaa tatnggtttg caccccaagc 420aaccaattta tnctgggcat agcgtnctta tggcnnggat ccttgccggg aaggggtgaa 480agcacttggc taaagagatg cattcctcna aaanctgaag gntaagtttg gacattngat 540gccggtt 5473675PRTOryza sativa 36Leu Met Ala Lys Leu Glu Lys Ala Asp Gln Ala Phe Cys Phe Thr Ser1 5 10 15Gly Met Ala Ala Leu Ala Ala Val Thr His Leu Leu Lys Ser Gly Gln20 25 30Glu Ile Val Ala Gly Glu Asp Ile Tyr Gly Gly Ser Asp Arg Leu Leu35 40 45Ser Gln Val Ala Pro Arg His Gly Ile Val Val Lys Arg Ile Asp Thr50 55 60Thr Lys Ile Ser Glu Val Thr Ser Ala Ile Gly65 70 75371733DNAGlycine max 37caaagacggc attgaagttg aacaatccat cactaacaca agcgcagaca acaacataac 60cctgctccaa acacatcaat ttcaataatg ttttcttctg caatttctca gaagcccttc 120cttcagtccc tcgtcattga tcgttacgct cagagcacaa ctgctgcaac caggtgggag 180tgcttggggt ttaacaagtc agaaaatttc agtaccaaga gagtgttgcg tgcagagggg 240ttcaagttga attgcttggt tgaaaataga gagatggaag tggagtcatc atcatcatct 300ttggtggatg atgctgccat gagcttaagt gaagaggatt taggggagcc tagtatttca 360acaatggtga tgaatttcga gagtaagttt gatccttttg gagcaattag taccccgctt 420taccaaacgg ctacttttaa gcagccttct gcaatagaaa atggtcccta tgactatacc 480agaagtggaa atcctactcg tgatgcttta gaaagtttac tagcaaagct tgataaagca 540gatagagccc tgtgcttcac cagtggaatg gctgctttga gtgctgttgt tcgtcttgtt 600ggaactggtg aggaaattgt caccggagat gatgtatatg gtggctcaga taggttgctg 660tctcaagtag ttccaaggac tggaattgtg gtgaaacggg taaatacatg tgatctagat 720gaggttgctg ctgccattgg actcaggact aagcttgtgt ggcttgagag tccaaccaat 780cctcggcttc aaatttctga tattcgaaaa atatcagaga tggctcattc acatggtgct 840cttgtgttag tggacaatag tataatgtca cctgtgttgt ctcagccatt ggaacttgga 900gcagatattg tcatgcactc agctacaaaa tttattgctg gacatagtga cattatggct 960ggtgtgcttg ctgtgaaggg tgaaaagttg ggaaaggaaa tgtatttctt gcaaaatgca 1020gagggttcag gcttagcacc atttgactgt tggctttgtt tgcgaggaat caagacaatg 1080gccctgcgaa ttgaaaagca acaggataac gcacagaaga ttgcagagtt ccttgcctcc 1140catcctcgag tgaaggaagt gaattatgct ggcttgcctg gtcatcctgg tcgtgattta 1200cactattctc aggcaaaggg tgcaggatct gtgcttagct tcttgactgg ttcattggca 1260ctttcaaagc atattgttga aactaccaaa tacttcagta taaccgtcag ctttgggagt 1320gtgaagtccc tcattagcat gccatgcttt atgtcacatg caagcatacc tgctgcagtt 1380cgcgaggcca gaggtttaac tgaagatctt gtacgaatat ctgtgggaat tgaggatgtg 1440aatgatctca ttgctgatct tggcaatgca cttagaactg gacctcttta atgtcttctc 1500caccccccca cccaaaaaga aaaaaattca tccttaagaa gttggattag catgttgagg 1560atttgggagc attgctatcc tgtctttgga ttcttgagag tggaaacttg aagtgttgct 1620tatgtgcatg taataaaatc aatatttcct gtaattttgt tgtaacaatt gttatcctta 1680ccttgcaata tcatgtcata caagttacta ttgaaaaaaa aaaaaaaaaa aaa 173338467PRTGlycine max 38Met Phe Ser Ser Ala Ile Ser Gln Lys Pro Phe Leu Gln Ser Leu Val1 5 10 15Ile Asp Arg Tyr Ala Gln Ser Thr Thr Ala Ala Thr Arg Trp Glu Cys20 25 30Leu Gly Phe Asn Lys Ser Glu Asn Phe Ser Thr Lys Arg Val Leu Arg35 40 45Ala Glu Gly Phe Lys Leu Asn Cys Leu Val Glu Asn Arg Glu Met Glu50 55 60Val Glu Ser Ser Ser Ser Ser Leu Val Asp Asp Ala Ala Met Ser Leu65 70 75 80Ser Glu Glu Asp Leu Gly Glu Pro Ser Ile Ser Thr Met Val Met Asn85 90 95Phe Glu Ser Lys Phe Asp Pro Phe Gly Ala Ile Ser Thr Pro Leu Tyr100 105 110Gln Thr Ala Thr Phe Lys Gln Pro Ser Ala Ile Glu Asn Gly Pro Tyr115 120 125Asp Tyr Thr Arg Ser Gly Asn Pro Thr Arg Asp Ala Leu Glu Ser Leu130 135 140Leu Ala Lys Leu Asp Lys Ala Asp Arg Ala Leu Cys Phe Thr Ser Gly145 150 155 160Met Ala Ala Leu Ser Ala Val Val Arg Leu Val Gly Thr Gly Glu Glu165 170 175Ile Val Thr Gly Asp Asp Val Tyr Gly Gly Ser Asp Arg Leu Leu Ser180 185 190Gln Val Val Pro Arg Thr Gly Ile Val Val Lys Arg Val Asn Thr Cys195 200 205Asp Leu Asp

Glu Val Ala Ala Ala Ile Gly Leu Arg Thr Lys Leu Val210 215 220Trp Leu Glu Ser Pro Thr Asn Pro Arg Leu Gln Ile Ser Asp Ile Arg225 230 235 240Lys Ile Ser Glu Met Ala His Ser His Gly Ala Leu Val Leu Val Asp245 250 255Asn Ser Ile Met Ser Pro Val Leu Ser Gln Pro Leu Glu Leu Gly Ala260 265 270Asp Ile Val Met His Ser Ala Thr Lys Phe Ile Ala Gly His Ser Asp275 280 285Ile Met Ala Gly Val Leu Ala Val Lys Gly Glu Lys Leu Gly Lys Glu290 295 300Met Tyr Phe Leu Gln Asn Ala Glu Gly Ser Gly Leu Ala Pro Phe Asp305 310 315 320Cys Trp Leu Cys Leu Arg Gly Ile Lys Thr Met Ala Leu Arg Ile Glu325 330 335Lys Gln Gln Asp Asn Ala Gln Lys Ile Ala Glu Phe Leu Ala Ser His340 345 350Pro Arg Val Lys Glu Val Asn Tyr Ala Gly Leu Pro Gly His Pro Gly355 360 365Arg Asp Leu His Tyr Ser Gln Ala Lys Gly Ala Gly Ser Val Leu Ser370 375 380Phe Leu Thr Gly Ser Leu Ala Leu Ser Lys His Ile Val Glu Thr Thr385 390 395 400Lys Tyr Phe Ser Ile Thr Val Ser Phe Gly Ser Val Lys Ser Leu Ile405 410 415Ser Met Pro Cys Phe Met Ser His Ala Ser Ile Pro Ala Ala Val Arg420 425 430Glu Ala Arg Gly Leu Thr Glu Asp Leu Val Arg Ile Ser Val Gly Ile435 440 445Glu Asp Val Asn Asp Leu Ile Ala Asp Leu Gly Asn Ala Leu Arg Thr450 455 460Gly Pro Leu46539637DNATriticum aestivummisc_feature(400)..(400)n is a, c, g, or t 39agcgtggcca cgatactgac cagcttcgag aactcgttcg acaagtatgg ggctctcagc 60acgccgctgt accagacggc caccttcaag cagccttcag caaccgttaa tggagcttat 120gattatacta gaagtggcaa ccctactcgt gatgttctcc agagccttat ggctaagctc 180gagaaggcag accaagcatt ctgcttcact agtgggatgg catcactggg ctgcagtaac 240acacctcctt caggctggac aagaaatagt tgctggagag gacatatatg gtggtctgat 300cgtctgctct cacaagttgt cccaagaaat ggaattgtag taaaacgggt cgatacaact 360aaaattaacg acgtgactgc tgcatcggac ccttgactan actagtttgg ttgaaancca 420caatcctcgt caacaattac tgtataagaa atctcaggga tactcatcca tggggactgg 480tttggnggca annttcatgt cccanggcta cctggccnat aaantggggn antatgggag 540catcagtaca aattatnctg gcnatgtcta ggtggatctc ntaaggggaa nttggnagga 600ttcttcaaaa cctagtnggt tgacttatgt ggttgtt 63740131PRTTriticum aestivummisc_feature(77)..(77)Xaa can be any naturally occurring amino acid 40Ser Val Ala Thr Ile Leu Thr Ser Phe Glu Asn Ser Phe Asp Lys Tyr1 5 10 15Gly Ala Leu Ser Thr Pro Leu Tyr Gln Thr Ala Thr Phe Lys Gln Pro20 25 30Ser Ala Thr Val Asn Gly Ala Tyr Asp Tyr Thr Arg Ser Gly Asn Pro35 40 45Thr Arg Asp Val Leu Gln Ser Leu Met Ala Lys Leu Glu Lys Ala Asp50 55 60Gln Ala Phe Cys Phe Thr Ser Gly Met Ala Ser Leu Xaa Ala Val Thr65 70 75 80His Leu Leu Gln Ala Gly Gln Glu Ile Val Ala Gly Glu Asp Ile Tyr85 90 95Gly Gly Xaa Asp Arg Leu Leu Ser Gln Val Val Pro Arg Asn Gly Ile100 105 110Val Val Lys Arg Val Asp Thr Thr Lys Ile Asn Asp Val Thr Ala Ala115 120 125Ser Asp Pro13041464PRTArabidopsis thaliana 41Met Thr Ser Ser Leu Ser Leu His Ser Ser Phe Val Pro Ser Phe Ala1 5 10 15Asp Leu Ser Asp Arg Gly Leu Ile Ser Lys Asn Ser Pro Thr Ser Val20 25 30Ser Ile Ser Lys Val Pro Thr Trp Glu Lys Lys Gln Ile Ser Asn Arg35 40 45Asn Ser Phe Lys Leu Asn Cys Val Met Glu Lys Ser Val Asp Gly Gln50 55 60Thr His Ser Thr Val Asn Asn Thr Thr Asp Ser Leu Asn Thr Met Asn65 70 75 80Ile Lys Glu Glu Ala Ser Val Ser Thr Leu Leu Val Asn Leu Asp Asn85 90 95Lys Phe Asp Pro Phe Asp Ala Met Ser Thr Pro Leu Tyr Gln Thr Ala100 105 110Thr Phe Lys Gln Pro Ser Ala Ile Glu Asn Gly Pro Tyr Asp Tyr Thr115 120 125Arg Ser Gly Asn Pro Thr Arg Asp Ala Leu Glu Ser Leu Leu Ala Lys130 135 140Leu Asp Lys Ala Asp Arg Ala Phe Cys Phe Thr Ser Gly Met Ala Ala145 150 155 160Leu Ser Ala Val Thr His Leu Ile Lys Asn Gly Glu Glu Ile Val Ala165 170 175Gly Asp Asp Val Tyr Gly Gly Ser Asp Arg Leu Leu Ser Gln Val Val180 185 190Pro Arg Ser Gly Val Val Val Lys Arg Val Asn Thr Thr Lys Leu Asp195 200 205Glu Val Ala Ala Ala Ile Gly Pro Gln Thr Lys Leu Val Trp Leu Glu210 215 220Ser Pro Thr Asn Pro Arg Gln Gln Ile Ser Asp Ile Arg Lys Ile Ser225 230 235 240Glu Met Ala His Ala Gln Gly Ala Leu Val Leu Val Asp Asn Ser Ile245 250 255Met Ser Pro Val Leu Ser Arg Pro Leu Glu Leu Gly Ala Asp Ile Val260 265 270Met His Ser Ala Thr Lys Phe Ile Ala Gly His Ser Asp Val Met Ala275 280 285Gly Val Leu Ala Val Lys Gly Glu Lys Leu Ala Lys Glu Val Tyr Phe290 295 300Leu Gln Asn Ser Glu Gly Ser Gly Leu Ala Pro Phe Asp Cys Trp Leu305 310 315 320Cys Leu Arg Gly Ile Lys Thr Met Ala Leu Arg Ile Glu Lys Gln Gln325 330 335Glu Asn Ala Arg Lys Ile Ala Met Tyr Leu Ser Ser His Pro Arg Val340 345 350Lys Lys Val Tyr Tyr Ala Gly Leu Pro Asp His Pro Gly His His Leu355 360 365His Phe Ser Gln Ala Lys Gly Ala Gly Ser Val Phe Ser Phe Ile Thr370 375 380Gly Ser Val Ala Leu Ser Lys His Leu Val Glu Thr Thr Lys Tyr Phe385 390 395 400Ser Ile Ala Val Ser Phe Gly Ser Val Lys Ser Leu Ile Ser Met Pro405 410 415Cys Phe Met Ser His Ala Ser Ile Pro Ala Glu Val Arg Glu Ala Arg420 425 430Gly Leu Thr Glu Asp Leu Val Arg Ile Ser Ala Gly Ile Glu Asp Val435 440 445Asp Asp Leu Ile Ser Asp Leu Asp Ile Ala Phe Lys Thr Phe Pro Leu450 455 460421113DNAZea mays 42gccgtccagg acctcgcggc ccctggggcg ttcgacggcg tcgacatcgc gctattcagc 60gccggcggga gcgtcagccg gaagtatggg cccgcggccg tcgccagcgg cgccgtagtt 120gtcgacaaca gctccgcgtt ccggatggag cccgaggtgc cgctcgtcat ccccgaggtc 180aaccccgagg ccatggcgaa cgtccgcctc gggcaggggg cgattgtggc aaatccgaat 240tgctcgacca tcatctgcct catggctgcc acgccgctcc atcgccacgc taaggtgtta 300aggatggttg tcagcacata ccaagcagca agtggtgcgg gtgctgcggc aatggaagaa 360ctcaagctgc agactcagga ggtcttggaa gggaaggcgc caacatgcaa cattttcaaa 420cagcagtatg cttttaatat attctcacac aatgcaccag ttcttgagaa tgggtataac 480gaggaggaaa tgaaaatggt gaaggagacc aggaaaattt ggaatgacaa ggaggtgaaa 540gtaactgcga cttgcatacg ggttcctgtg atgcgcgcac atgctgaaag tgtcaatcta 600cagtttgaaa agccacttga tgaggatact gcaagagaaa ttttgagagc agctcctggt 660gttaccatta ttgatgaccg agcttccaat cgctttccta cacctctgga ggtatcagac 720aaagatgacg tagcagtggg taggattcgt caggacttgt ccctggatgg taaccgaggg 780ttggacatat ttgtgtgtgg tgatcagata cgtaaaggcg ccgcactcaa tgccgttcag 840attgctgaaa tgctgctgaa gtgaatgtga cctaaccctc ttgtccctcc ctccctgtcc 900ctaattgctc tgatcaaatg ctggactgta ctctgattag tttgtcctca attttggtcg 960cctgttctgt attctgccgt gctagtgcaa taattgtgtt atgggcttga gttatctgct 1020gtacgcataa gtgggctcct aaactgggaa ataatgggcc gtccttattc agcattccgg 1080tttatatctt gttcaaaaaa aaaaaaaaaa ata 111343287PRTZea mays 43Ala Val Gln Asp Leu Ala Ala Pro Gly Ala Phe Asp Gly Val Asp Ile1 5 10 15Ala Leu Phe Ser Ala Gly Gly Ser Val Ser Arg Lys Tyr Gly Pro Ala20 25 30Ala Val Ala Ser Gly Ala Val Val Val Asp Asn Ser Ser Ala Phe Arg35 40 45Met Glu Pro Glu Val Pro Leu Val Ile Pro Glu Val Asn Pro Glu Ala50 55 60Met Ala Asn Val Arg Leu Gly Gln Gly Ala Ile Val Ala Asn Pro Asn65 70 75 80Cys Ser Thr Ile Ile Cys Leu Met Ala Ala Thr Pro Leu His Arg His85 90 95Ala Lys Val Leu Arg Met Val Val Ser Thr Tyr Gln Ala Ala Ser Gly100 105 110Ala Gly Ala Ala Ala Met Glu Glu Leu Lys Leu Gln Thr Gln Glu Val115 120 125Leu Glu Gly Lys Ala Pro Thr Cys Asn Ile Phe Lys Gln Gln Tyr Ala130 135 140Phe Asn Ile Phe Ser His Asn Ala Pro Val Leu Glu Asn Gly Tyr Asn145 150 155 160Glu Glu Glu Met Lys Met Val Lys Glu Thr Arg Lys Ile Trp Asn Asp165 170 175Lys Glu Val Lys Val Thr Ala Thr Cys Ile Arg Val Pro Val Met Arg180 185 190Ala His Ala Glu Ser Val Asn Leu Gln Phe Glu Lys Pro Leu Asp Glu195 200 205Asp Thr Ala Arg Glu Ile Leu Arg Ala Ala Pro Gly Val Thr Ile Ile210 215 220Asp Asp Arg Ala Ser Asn Arg Phe Pro Thr Pro Leu Glu Val Ser Asp225 230 235 240Lys Asp Asp Val Ala Val Gly Arg Ile Arg Gln Asp Leu Ser Leu Asp245 250 255Gly Asn Arg Gly Leu Asp Ile Phe Val Cys Gly Asp Gln Ile Arg Lys260 265 270Gly Ala Ala Leu Asn Ala Val Gln Ile Ala Glu Met Leu Leu Lys275 280 285441402DNAOryza sativa 44gcccaactcc caaaacccta gaaccgcgcc gccacaatgc aggccgccgc cgccgccgtc 60caccgcccgc acctcctcgg cgcctacccc ggcggtggcc gcgcgcgccg cccgtcgtcc 120accgtgcgga tggcgcttcg ggaggacggg ccgtcggtgg cgatcgtggg cgcgacgggc 180gccgtcggcc aggagttcct ccgcgtcatc tcctcccggg gcttccccta ccggagcctc 240cgcctcctcg ccagcgagcg ctccgcgggg aagcgcctcc cgttcgaggg ccaggagtac 300accgtccagg acctcgccgc gccgggcgcg ttcgacgggg tggacatcgc gctcttcagc 360gccggcggcg gggtcagccg cgcccacgct cccgcggccg tcgccagcgg cgccgtcgtc 420gtggacaaca gctccgcctt ccggatggac cccgaggtgc cgctcgtcat ccccgaggtc 480aatcccgagg ccatggcgca cgtccggctg ggaaaggggg ctattgtggc caacccgaac 540tgttccacca tcatctgcct catggctgcc acacctctgc accgccacgc caaggtggta 600aggatggttg tcagcactta ccaagcagca agtggtgctg gggctgcggc catggaagaa 660ctcaaacttc aaactcaaga ggtcttggcg gggaaagcac caacatgcaa cattttcagt 720cagcagtatg cttttaatat attttcacat aatgcaccaa ttgttgaaaa tgggtacaat 780gaggaggaga tgaagatggt gaaggagacc agaaaaatct ggaatgataa agatgtgaag 840gtaactgcaa cctgcatacg agttcctgtg atgcgtgcac atgctgaaag tgtgaatcta 900cagtttgaaa agccacttga tgaggatact gcaagggaaa tcttgagggc agctgaaggt 960gttaccatta ttgatgaccg tgcttccaat cgcttcccca cacctcttga ggtatcggat 1020aaagatgatg tagcagtggg tagaattcgt caggatttgt cgcaagatga taacaaaggg 1080ctggacatat ttgtttgtgg agatcaaata cgtaaaggtg ctgcactcaa tgctgtgcag 1140attgctgaaa tgctactcaa gtgattttct tttctgtacc tttctctcct tgcccctctt 1200tgctctagtc attgtttgac ggatgtactc tggttagtat gagatcaatt ttgatcatct 1260tttgtaatct atattcctag tgaaataaat gtaaaacggt tttgctctat cttctgcaca 1320agtgtagaag aaatctgaaa ttgggaaatt ggagtgtggc ccttgttcaa aaaaaaaaaa 1380aaaaaaaaaa aaaaaaaaaa aa 140245375PRTOryza sativa 45Met Gln Ala Ala Ala Ala Ala Val His Arg Pro His Leu Leu Gly Ala1 5 10 15Tyr Pro Gly Gly Gly Arg Ala Arg Arg Pro Ser Ser Thr Val Arg Met20 25 30Ala Leu Arg Glu Asp Gly Pro Ser Val Ala Ile Val Gly Ala Thr Gly35 40 45Ala Val Gly Gln Glu Phe Leu Arg Val Ile Ser Ser Arg Gly Phe Pro50 55 60Tyr Arg Ser Leu Arg Leu Leu Ala Ser Glu Arg Ser Ala Gly Lys Arg65 70 75 80Leu Pro Phe Glu Gly Gln Glu Tyr Thr Val Gln Asp Leu Ala Ala Pro85 90 95Gly Ala Phe Asp Gly Val Asp Ile Ala Leu Phe Ser Ala Gly Gly Gly100 105 110Val Ser Arg Ala His Ala Pro Ala Ala Val Ala Ser Gly Ala Val Val115 120 125Val Asp Asn Ser Ser Ala Phe Arg Met Asp Pro Glu Val Pro Leu Val130 135 140Ile Pro Glu Val Asn Pro Glu Ala Met Ala His Val Arg Leu Gly Lys145 150 155 160Gly Ala Ile Val Ala Asn Pro Asn Cys Ser Thr Ile Ile Cys Leu Met165 170 175Ala Ala Thr Pro Leu His Arg His Ala Lys Val Val Arg Met Val Val180 185 190Ser Thr Tyr Gln Ala Ala Ser Gly Ala Gly Ala Ala Ala Met Glu Glu195 200 205Leu Lys Leu Gln Thr Gln Glu Val Leu Ala Gly Lys Ala Pro Thr Cys210 215 220Asn Ile Phe Ser Gln Gln Tyr Ala Phe Asn Ile Phe Ser His Asn Ala225 230 235 240Pro Ile Val Glu Asn Gly Tyr Asn Glu Glu Glu Met Lys Met Val Lys245 250 255Glu Thr Arg Lys Ile Trp Asn Asp Lys Asp Val Lys Val Thr Ala Thr260 265 270Cys Ile Arg Val Pro Val Met Arg Ala His Ala Glu Ser Val Asn Leu275 280 285Gln Phe Glu Lys Pro Leu Asp Glu Asp Thr Ala Arg Glu Ile Leu Arg290 295 300Ala Ala Glu Gly Val Thr Ile Ile Asp Asp Arg Ala Ser Asn Arg Phe305 310 315 320Pro Thr Pro Leu Glu Val Ser Asp Lys Asp Asp Val Ala Val Gly Arg325 330 335Ile Arg Gln Asp Leu Ser Gln Asp Asp Asn Lys Gly Leu Asp Ile Phe340 345 350Val Cys Gly Asp Gln Ile Arg Lys Gly Ala Ala Leu Asn Ala Val Gln355 360 365Ile Ala Glu Met Leu Leu Lys370 375461391DNAGlycine max 46gcacgagctt cactctctgt tttgcgccac aaccacctct tctcgggccc cctcccggcc 60cgccccaagc ccacctcctc ctcctcctcc aggatccgaa tgtccctccg cgagaacggc 120ccctccatcg ccgtcgtggg cgtcaccggc gccgtcggcc aggagttcct ctccgtcctc 180tccgaccgcg acttccccta ccgctccatt catatgctgg cttccaagcg ctccgctggc 240cgccgcatca ccttcgagga cagggactac gtcgtccagg agctcacgcc ggagagcttc 300gacggtgtcg acatcgcgct cttcagcgcc ggcggctcca tcagcaagca cttcggcccc 360atcgccgtca atcgtggaac ggtcgtggtc gacaacagct ccgcgtttcg gatgaacgag 420aaggtgcctt tggtaattcc cgaagtgaac cccgaagcaa tgcaaaacat caaagccgga 480acgggaaagg gcgcactcat tgctaaccct aattgctcca ccattatatg cttgatggct 540gctacccctc ttcatcgacg tgccaaggtg ttacgtatgg ttgttagtac ctatcaggct 600gcgagtggtg ctggtgctgc tgcaatggaa gagcttgagc tgcaaactcg tgaggtgttg 660gaaggaaaac cacccacttg taaaatattt aaccgacagt atgcttttaa tctattctca 720cataatgcgt ctgttctttc aaatggatat aatgaagaag aaatgaaaat ggtcaaggag 780accaggaaaa tctggaatga caaggatgtt aaagtaactg ccacatgcat acgagttccc 840atcatgcgag ctcatgctga gagtgtgaat cttcaatttg aaagacccct tgatgaggac 900actgcaagag atattctgaa aaatgctcca ggtgtagtgg ttattgatga tcgtgaatcc 960aatcattttc ctactccact ggaagtgtca aacaaggatg atgttgctgt tggtaggatt 1020cggcaggacc tgtctcagga tgggaatcaa gggttggaca tctttgtatg tggggatcaa 1080attcgcaagg gagctgcact taacgcaatc cagattgctg agatgttgct atgagttctg 1140gtttttcaag gatctggtac ttaaagatta tgcttctttt gaaacagttt tgtatgtgct 1200agttgtatgt ggttattcat ttcttttgtg atgtttaact agtccaagta tcttttcaac 1260gatgtggtag cacactagct ggaaacagtt tttttaaggt cttggtgcgt aatatctgca 1320atccttttca ccgggaataa caagcactgg ttatggcaaa aaaaaaaaaa aaaaaaaaaa 1380aaaaaaaaaa a 139147377PRTGlycine max 47Ala Arg Ala Ser Leu Ser Val Leu Arg His Asn His Leu Phe Ser Gly1 5 10 15Pro Leu Pro Ala Arg Pro Lys Pro Thr Ser Ser Ser Ser Ser Arg Ile20 25 30Arg Met Ser Leu Arg Glu Asn Gly Pro Ser Ile Ala Val Val Gly Val35 40 45Thr Gly Ala Val Gly Gln Glu Phe Leu Ser Val Leu Ser Asp Arg Asp50 55 60Phe Pro Tyr Arg Ser Ile His Met Leu Ala Ser Lys Arg Ser Ala Gly65 70 75 80Arg Arg Ile Thr Phe Glu Asp Arg Asp Tyr Val Val Gln Glu Leu Thr85 90 95Pro Glu Ser Phe Asp Gly Val Asp Ile Ala Leu Phe Ser Ala Gly Gly100 105 110Ser Ile Ser Lys His Phe Gly Pro Ile Ala Val Asn Arg Gly Thr Val115 120 125Val Val Asp Asn Ser Ser Ala Phe Arg Met Asn Glu Lys Val Pro Leu130 135 140Val Ile Pro Glu Val Asn Pro Glu Ala Met Gln Asn Ile Lys Ala Gly145 150 155 160Thr Gly Lys Gly Ala Leu Ile Ala Asn Pro Asn Cys Ser Thr Ile Ile165 170 175Cys Leu Met Ala Ala Thr Pro Leu His Arg Arg Ala Lys Val Leu Arg180 185 190Met Val Val Ser Thr Tyr Gln Ala Ala Ser Gly Ala Gly Ala Ala Ala195 200 205Met Glu Glu Leu Glu Leu Gln Thr Arg Glu Val Leu Glu Gly Lys Pro210 215 220Pro Thr Cys Lys Ile Phe Asn Arg Gln Tyr Ala Phe Asn Leu Phe Ser225 230 235 240His Asn Ala Ser Val Leu Ser Asn Gly Tyr Asn Glu Glu Glu Met Lys245 250 255Met Val Lys Glu Thr Arg Lys Ile Trp Asn Asp Lys Asp Val Lys Val260 265 270Thr Ala Thr Cys Ile Arg Val

Pro Ile Met Arg Ala His Ala Glu Ser275 280 285Val Asn Leu Gln Phe Glu Arg Pro Leu Asp Glu Asp Thr Ala Arg Asp290 295 300Ile Leu Lys Asn Ala Pro Gly Val Val Val Ile Asp Asp Arg Glu Ser305 310 315 320Asn His Phe Pro Thr Pro Leu Glu Val Ser Asn Lys Asp Asp Val Ala325 330 335Val Gly Arg Ile Arg Gln Asp Leu Ser Gln Asp Gly Asn Gln Gly Leu340 345 350Asp Ile Phe Val Cys Gly Asp Gln Ile Arg Lys Gly Ala Ala Leu Asn355 360 365Ala Ile Gln Ile Ala Glu Met Leu Leu370 375481470DNAGlycine max 48gcacgaggtc tgttttaaaa tccaacactt aatctctctc ttcgcagcct aaaatcccaa 60tggcttcact ctctgttttg cgccacaacc acctcttctc gggccccctc ccggcccgcc 120ccaagcccac ctcctcctcc tcctccagga tccgaatgtc cctccgcgag aacggcccct 180ccatcgccgt cgtgggcgtc accggcgccg tcggccagga gttcctctcc gtcctctccg 240accgcgactt cccctaccgc tccattcata tgctggcttc caagcgctcc gctggccgcc 300gcatcacctt cgaggacagg gactacgtcg tccaggagct cacgccggag agcttcgacg 360gtgtcgacat cgcgctcttc agcgccggcg gctccatcag caagcacttc ggccccatcg 420ccgtcaatcg tggaacggtc gtggtcgaca acagctccgc gtttcggatg gacgagaagg 480tgcctttggt aattcccgaa gtgaaccccg aagcaatgca aaacatcaaa gccggaacgg 540gaaagggcgc actcattgct aaccctaatt gctccaccat tagatgcttg aaggctgcta 600cccctcttca tcgacgtgcc aaggtgttac gtatggttgt tagtacctat caggctgcga 660gtggtgctgg tgctgctgca atggaagagc ttgagctgca aactcgtgag gtgttggaag 720gaaaaccacc cacttgtaaa atatttaacc gacagtatgc ttttaatcta ttctcacata 780atgcgtctgt tctttcaaat ggatataatg aagaagaaat gaaaatggtc aaggagacca 840ggaaaatctg gaatgacaag gatgttaaag taactgccac atgcatacga gttcccatca 900tgcgagctca tgctgagagt gtgaatcttc aatttgaaag accccttgat gaggacactg 960caagagatat tctgaaaaat gctccaggtg tagtggttat tgatgatcgt gaatccaatc 1020attttcctac tccactggaa gtgtcaaaca aggatgatgt tgctgttggt aggattcggc 1080aggacctgtc tcaggatggg aatcaagggt tggacatctt tgtatgtggg gatcaaattc 1140gcaagggagc tgcacttaac gcaatccaga ttgctgagat gttgctatga gttctggttt 1200ttcaaggatc tggtacttaa agattatgct tcttttgaaa cagttttgta tgtgctagtt 1260gtatgtggtt attcatttct tttgtgatgt ttaactagtc caagtatctt ttcaacgatg 1320tggtagcaca ctagctggaa acagtttttt taaggtcttg gtgcgtaata tctgcaatcc 1380ttttcaccgg gaataacaag cactggtttt ggcaaaaaaa aaaaaaaaaa aaaaaaaaaa 1440aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa 147049376PRTGlycine max 49Met Ala Ser Leu Ser Val Leu Arg His Asn His Leu Phe Ser Gly Pro1 5 10 15Leu Pro Ala Arg Pro Lys Pro Thr Ser Ser Ser Ser Ser Arg Ile Arg20 25 30Met Ser Leu Arg Glu Asn Gly Pro Ser Ile Ala Val Val Gly Val Thr35 40 45Gly Ala Val Gly Gln Glu Phe Leu Ser Val Leu Ser Asp Arg Asp Phe50 55 60Pro Tyr Arg Ser Ile His Met Leu Ala Ser Lys Arg Ser Ala Gly Arg65 70 75 80Arg Ile Thr Phe Glu Asp Arg Asp Tyr Val Val Gln Glu Leu Thr Pro85 90 95Glu Ser Phe Asp Gly Val Asp Ile Ala Leu Phe Ser Ala Gly Gly Ser100 105 110Ile Ser Lys His Phe Gly Pro Ile Ala Val Asn Arg Gly Thr Val Val115 120 125Val Asp Asn Ser Ser Ala Phe Arg Met Asp Glu Lys Val Pro Leu Val130 135 140Ile Pro Glu Val Asn Pro Glu Ala Met Gln Asn Ile Lys Ala Gly Thr145 150 155 160Gly Lys Gly Ala Leu Ile Ala Asn Pro Asn Cys Ser Thr Ile Arg Cys165 170 175Leu Lys Ala Ala Thr Pro Leu His Arg Arg Ala Lys Val Leu Arg Met180 185 190Val Val Ser Thr Tyr Gln Ala Ala Ser Gly Ala Gly Ala Ala Ala Met195 200 205Glu Glu Leu Glu Leu Gln Thr Arg Glu Val Leu Glu Gly Lys Pro Pro210 215 220Thr Cys Lys Ile Phe Asn Arg Gln Tyr Ala Phe Asn Leu Phe Ser His225 230 235 240Asn Ala Ser Val Leu Ser Asn Gly Tyr Asn Glu Glu Glu Met Lys Met245 250 255Val Lys Glu Thr Arg Lys Ile Trp Asn Asp Lys Asp Val Lys Val Thr260 265 270Ala Thr Cys Ile Arg Val Pro Ile Met Arg Ala His Ala Glu Ser Val275 280 285Asn Leu Gln Phe Glu Arg Pro Leu Asp Glu Asp Thr Ala Arg Asp Ile290 295 300Leu Lys Asn Ala Pro Gly Val Val Val Ile Asp Asp Arg Glu Ser Asn305 310 315 320His Phe Pro Thr Pro Leu Glu Val Ser Asn Lys Asp Asp Val Ala Val325 330 335Gly Arg Ile Arg Gln Asp Leu Ser Gln Asp Gly Asn Gln Gly Leu Asp340 345 350Ile Phe Val Cys Gly Asp Gln Ile Arg Lys Gly Ala Ala Leu Asn Ala355 360 365Ile Gln Ile Ala Glu Met Leu Leu370 375501609DNATriticum aestivum 50caccaccacc cacctaccca aatcccagcc gccctaaaac cctaggccgc caaacccgcc 60gccgccgccg ccgcaatgca ggccgccgca gccgtccacc ggccacacct cctcgcggcg 120tccccgctcg ggggccgcgc cagccgccgg ccctccacgg tccgcatggc gctccgcgag 180gacgggccct ccgtggccat cgtgggcgcc accggcgcgg tggggcagga gttcctccgc 240gtcatcaccg cccgcgactt cccctaccgc agcctgcgcc tcctcgccag cgagcgctcc 300gcgggcaagc gcatcgactt cgagggccgg gactacaccg tccaggacct cgcggcgccg 360ggggccttcg acggggtcga catcgcgctc ttcagcgccg gcgggagcat cagccgcgcc 420cacgcgcccg ccgccgtcgc cagcggcgcc gtcgtcgtgg ataacagctc cgcctaccgg 480atggaccccg acgtgccgct cgtcatcccg gaggttaacc ccgaggccat ggccgacgtc 540cggctcggga aaggggctat tgtggccaac cccaactgtt ccaccatcat ctgcctcatg 600gctgtcacgc cgctgcatcg ccacgccaag gtgaaaagga tggttgtcag cacataccaa 660gcagcaagtg gtgctggtgc tgcagccatg gaagaactca aacttcagac tcgagaggtc 720ttggaaggaa agccaccaac ctgtaacatt ttcagtcaac agtatgcttt taatatattt 780tcgcataatg cacctattgt tgaaaatggc tataatgagg aagagatgaa aatggtgaag 840gagaccagaa aaatctggaa tgacaaggat gtaagagtaa ctgcaacttg tatacgggtt 900cctacgatgc gcgcgcatgc cgaaagcgtg aatctacagt ttgaaaagcc acttgatgag 960gacactgcca gagaaatctt gagggcagct cctggtgtta ccattagtga cgaccgtgct 1020gccaaccgct tccctacacc actggaggta tcggataaag atgacgtatc agttggtagg 1080attcgccagg acttgtcaca agatgataac agagggttgg agttatttgt ctgtggagac 1140cagatacgta aaggcgccgc gctgaacgct gtgcagattg ctgaaatgct actgaagtga 1200ccgccttttt accattgtct catgtgccac gttgctctat ccattgatgg attgatgtac 1260tctagtcact ttcaacccag ttttggtcgt cgtctttttt gtaatctgtc aacctagcag 1320aagaagtgta agacgggctt tagtcatctg ttgcacacaa aagtgcagcc acaagtttag 1380aaaaggaggg ttttcacttg ttcggatttt gccttaggtt ggactttgtt gcaagtttgt 1440cgtttgtttc ttgaaagctg gtctgctgta actttacccc caaagccctc gagataacga 1500ggcgtcctgt ggggacctaa aaaaaaaaaa aaaaaaaaaa aaaaaacccc aaaaaaaaaa 1560aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaa 160951374PRTTriticum aestivum 51Met Gln Ala Ala Ala Ala Val His Arg Pro His Leu Leu Ala Ala Ser1 5 10 15Pro Leu Gly Gly Arg Ala Ser Arg Arg Pro Ser Thr Val Arg Met Ala20 25 30Leu Arg Glu Asp Gly Pro Ser Val Ala Ile Val Gly Ala Thr Gly Ala35 40 45Val Gly Gln Glu Phe Leu Arg Val Ile Thr Ala Arg Asp Phe Pro Tyr50 55 60Arg Ser Leu Arg Leu Leu Ala Ser Glu Arg Ser Ala Gly Lys Arg Ile65 70 75 80Asp Phe Glu Gly Arg Asp Tyr Thr Val Gln Asp Leu Ala Ala Pro Gly85 90 95Ala Phe Asp Gly Val Asp Ile Ala Leu Phe Ser Ala Gly Gly Ser Ile100 105 110Ser Arg Ala His Ala Pro Ala Ala Val Ala Ser Gly Ala Val Val Val115 120 125Asp Asn Ser Ser Ala Tyr Arg Met Asp Pro Asp Val Pro Leu Val Ile130 135 140Pro Glu Val Asn Pro Glu Ala Met Ala Asp Val Arg Leu Gly Lys Gly145 150 155 160Ala Ile Val Ala Asn Pro Asn Cys Ser Thr Ile Ile Cys Leu Met Ala165 170 175Val Thr Pro Leu His Arg His Ala Lys Val Lys Arg Met Val Val Ser180 185 190Thr Tyr Gln Ala Ala Ser Gly Ala Gly Ala Ala Ala Met Glu Glu Leu195 200 205Lys Leu Gln Thr Arg Glu Val Leu Glu Gly Lys Pro Pro Thr Cys Asn210 215 220Ile Phe Ser Gln Gln Tyr Ala Phe Asn Ile Phe Ser His Asn Ala Pro225 230 235 240Ile Val Glu Asn Gly Tyr Asn Glu Glu Glu Met Lys Met Val Lys Glu245 250 255Thr Arg Lys Ile Trp Asn Asp Lys Asp Val Arg Val Thr Ala Thr Cys260 265 270Ile Arg Val Pro Thr Met Arg Ala His Ala Glu Ser Val Asn Leu Gln275 280 285Phe Glu Lys Pro Leu Asp Glu Asp Thr Ala Arg Glu Ile Leu Arg Ala290 295 300Ala Pro Gly Val Thr Ile Ser Asp Asp Arg Ala Ala Asn Arg Phe Pro305 310 315 320Thr Pro Leu Glu Val Ser Asp Lys Asp Asp Val Ser Val Gly Arg Ile325 330 335Arg Gln Asp Leu Ser Gln Asp Asp Asn Arg Gly Leu Glu Leu Phe Val340 345 350Cys Gly Asp Gln Ile Arg Lys Gly Ala Ala Leu Asn Ala Val Gln Ile355 360 365Ala Glu Met Leu Leu Lys37052340PRTAquifex aeolicus 52Met Gly Tyr Arg Val Ala Ile Val Gly Ala Thr Gly Glu Val Gly Arg1 5 10 15Thr Phe Leu Lys Val Leu Glu Glu Arg Asn Phe Pro Val Asp Glu Leu20 25 30Val Leu Tyr Ala Ser Glu Arg Ser Glu Gly Lys Val Leu Thr Phe Lys35 40 45Gly Lys Glu Tyr Thr Val Lys Ala Leu Asn Lys Glu Asn Ser Phe Lys50 55 60Gly Ile Asp Ile Ala Leu Phe Ser Ala Gly Gly Ser Thr Ser Lys Glu65 70 75 80Trp Ala Pro Lys Phe Ala Lys Asp Gly Val Val Val Ile Asp Asn Ser85 90 95Ser Ala Trp Arg Met Asp Pro Asp Val Pro Leu Val Val Pro Glu Val100 105 110Asn Pro Glu Asp Val Lys Asp Phe Lys Lys Lys Gly Ile Ile Ala Asn115 120 125Pro Asn Cys Ser Thr Ile Gln Met Val Val Ala Leu Lys Pro Ile Tyr130 135 140Asp Lys Ala Gly Ile Lys Arg Val Val Val Ser Thr Tyr Gln Ala Val145 150 155 160Ser Gly Ala Gly Ala Lys Ala Ile Glu Asp Leu Lys Asn Gln Thr Lys165 170 175Ala Trp Cys Glu Gly Lys Glu Met Pro Lys Ala Gln Lys Phe Pro His180 185 190Gln Ile Ala Phe Asn Ala Leu Pro His Ile Asp Val Phe Phe Glu Asp195 200 205Gly Tyr Thr Lys Glu Glu Asn Lys Met Leu Tyr Glu Thr Arg Lys Ile210 215 220Met His Asp Glu Asn Ile Lys Val Ser Ala Thr Cys Val Arg Ile Pro225 230 235 240Val Phe Tyr Gly His Ser Glu Ser Ile Ser Met Glu Thr Glu Lys Glu245 250 255Ile Ser Pro Glu Glu Ala Arg Glu Val Leu Lys Asn Ala Pro Gly Val260 265 270Ile Val Ile Asp Asn Pro Gln Asn Asn Glu Tyr Pro Met Pro Ile Met275 280 285Ala Glu Gly Arg Asp Glu Val Phe Val Gly Arg Ile Arg Lys Asp Arg290 295 300Val Phe Glu Pro Gly Leu Ser Met Trp Val Val Ala Asp Asn Ile Arg305 310 315 320Lys Gly Ala Ala Thr Asn Ala Val Gln Ile Ala Glu Leu Leu Val Lys325 330 335Glu Gly Leu Ile340531727DNAGlycine max 53ttgcaacaca cattgtcttg tcggcaaaat cttccaccaa caacacacag ccatggcagg 60ctcaaacatt ctttctcact ctccttccct tcccaaaacc tacagccact ccttaaacca 120aaacgcgtta tcccaaaagc ttttttttct gcccctcaaa ttcaaagcca ccacaaaacc 180acgtgctctc agagcggttc tctcgcagaa cgctgtcaaa acctcggtgg aggacacaaa 240gaacgctcat tttcagcact gtttcaccaa atccgaagat gggtatctgt actgtgaggg 300cctcaaggtg catgacatca tggaatctgt tgagagaaga cctttctatt tgtacagcaa 360gccccagata actaggaatg ttgaagccta caaggatgca ttggaagggt tgaactccat 420aattggttat gccattaagg ccaataataa cttgaagatt ttggaacatt tgaggcactt 480gggttgtggt gctgtgcttg ttagtgggaa tgagctgaag ttggctcttc gagctggctt 540tgatcccaca aggtgtatct ttaatgggaa tgggaaaatc ttggaggatt tggtcttggc 600tgctcaggaa ggtgtgtttg tcaacattga tagtgagttt gacttggaaa acattgtaga 660ggctgcaaaa agggctggga agaaggtcaa tgttttactt cggattaatc ctgatgtgga 720tccacaggtt catccttatg ttgccactgg gaataagaac tctaaatttg gcattagaaa 780tgagaagctg cagtgctttt tagatgcagt gaaggaacat cctaatgagc tcaaacttgt 840aggggcccac tgccatcttg gttcaacaat taccaaggtt gacattttca gggatgcagc 900caccattatg atcaactaca ttgaccaaat ccgagatcag ggttttgaag ttgattactt 960aaatattggt ggaggacttg ggatagatta ttatcattct ggtgccatcc ttcctacacc 1020tagagatctc attgacactg tacgagatct tgttatttca cgtggtctta atctcatcat 1080tgaaccagga agatcactca ttgcaaacac gtgttgctta gttaaccggg tgacaggtgt 1140taaaactaat ggatctaaaa acttcattgt aattgatgga agtatggctg aacttatccg 1200ccctagtctt tatgatgctt accagcatat agagctggtt tcccctgccc cgtcaaatgc 1260tgaaacagaa acttttgatg tggttggccc tgtctgtgag tctgcagatt tcttaggaaa 1320aggaagagaa cttcctactc cagccaaggg tactggtttg gttgttcatg atgctggtgc 1380ttattgcatg agcatggcat caacctacaa tctaaagatg cggcctcctg agtattgggt 1440tgaagatgat ggatcagtga gcaaaataag acatggagag acttttgaag accacattcg 1500gttttttgag gggctttgag ctaataattt atcttgtagg aaagaaggct ggagaattgt 1560tatgtacttg gagtttgaat ctttcctcgt caatgaatgc atgactcttg tagttctgtt 1620tcttccgttc taattgaatg ttgactccca tgacaggaac agagaataaa gttgatttca 1680gttaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaa 172754505PRTGlycine max 54Cys Asn Thr His Cys Leu Val Gly Lys Ile Phe His Gln Gln His Thr1 5 10 15Ala Met Ala Gly Ser Asn Ile Leu Ser His Ser Pro Ser Leu Pro Lys20 25 30Thr Tyr Ser His Ser Leu Asn Gln Asn Ala Leu Ser Gln Lys Leu Phe35 40 45Phe Leu Pro Leu Lys Phe Lys Ala Thr Thr Lys Pro Arg Ala Leu Arg50 55 60Ala Val Leu Ser Gln Asn Ala Val Lys Thr Ser Val Glu Asp Thr Lys65 70 75 80Asn Ala His Phe Gln His Cys Phe Thr Lys Ser Glu Asp Gly Tyr Leu85 90 95Tyr Cys Glu Gly Leu Lys Val His Asp Ile Met Glu Ser Val Glu Arg100 105 110Arg Pro Phe Tyr Leu Tyr Ser Lys Pro Gln Ile Thr Arg Asn Val Glu115 120 125Ala Tyr Lys Asp Ala Leu Glu Gly Leu Asn Ser Ile Ile Gly Tyr Ala130 135 140Ile Lys Ala Asn Asn Asn Leu Lys Ile Leu Glu His Leu Arg His Leu145 150 155 160Gly Cys Gly Ala Val Leu Val Ser Gly Asn Glu Leu Lys Leu Ala Leu165 170 175Arg Ala Gly Phe Asp Pro Thr Arg Cys Ile Phe Asn Gly Asn Gly Lys180 185 190Ile Leu Glu Asp Leu Val Leu Ala Ala Gln Glu Gly Val Phe Val Asn195 200 205Ile Asp Ser Glu Phe Asp Leu Glu Asn Ile Val Glu Ala Ala Lys Arg210 215 220Ala Gly Lys Lys Val Asn Val Leu Leu Arg Ile Asn Pro Asp Val Asp225 230 235 240Pro Gln Val His Pro Tyr Val Ala Thr Gly Asn Lys Asn Ser Lys Phe245 250 255Gly Ile Arg Asn Glu Lys Leu Gln Cys Phe Leu Asp Ala Val Lys Glu260 265 270His Pro Asn Glu Leu Lys Leu Val Gly Ala His Cys His Leu Gly Ser275 280 285Thr Ile Thr Lys Val Asp Ile Phe Arg Asp Ala Ala Thr Ile Met Ile290 295 300Asn Tyr Ile Asp Gln Ile Arg Asp Gln Gly Phe Glu Val Asp Tyr Leu305 310 315 320Asn Ile Gly Gly Gly Leu Gly Ile Asp Tyr Tyr His Ser Gly Ala Ile325 330 335Leu Pro Thr Pro Arg Asp Leu Ile Asp Thr Val Arg Asp Leu Val Ile340 345 350Ser Arg Gly Leu Asn Leu Ile Ile Glu Pro Gly Arg Ser Leu Ile Ala355 360 365Asn Thr Cys Cys Leu Val Asn Arg Val Thr Gly Val Lys Thr Asn Gly370 375 380Ser Lys Asn Phe Ile Val Ile Asp Gly Ser Met Ala Glu Leu Ile Arg385 390 395 400Pro Ser Leu Tyr Asp Ala Tyr Gln His Ile Glu Leu Val Ser Pro Ala405 410 415Pro Ser Asn Ala Glu Thr Glu Thr Phe Asp Val Val Gly Pro Val Cys420 425 430Glu Ser Ala Asp Phe Leu Gly Lys Gly Arg Glu Leu Pro Thr Pro Ala435 440 445Lys Gly Thr Gly Leu Val Val His Asp Ala Gly Ala Tyr Cys Met Ser450 455 460Met Ala Ser Thr Tyr Asn Leu Lys Met Arg Pro Pro Glu Tyr Trp Val465 470 475 480Glu Asp Asp Gly Ser Val Ser Lys Ile Arg His Gly Glu Thr Phe Glu485 490 495Asp His Ile Arg Phe Phe Glu Gly Leu500 50555858DNATriticum aestivum 55tttgagttgg agtacctgaa tattggaggt ggtttgggga tagactacca ccacactggt 60gcagtcttgc ctacacctat ggatcttatc aacactgtcc gggaattggt cctctcacgg 120gatcttactc tcattattga acctggaaga tccctgatcg ccaatacttg ctgcttcgtc 180aataaggtca ctggtgtaaa atcgaatggc acgaagaatt tcattgtagt tgatggcagc 240atggccgagc tcatcaggcc tagtctatat ggagcatatc agcatataga actagtttct 300ccctctccag gtgcagaagt agcaaccttc gatattgttg ggccagtctg cgaatctgca 360gatttccttg gcaaagacag ggagcttcca acacctgaca agggagctgg tttggttgtc 420cacgacgcag

gagcctactg catgagcatg gcttcgacct acaacctgaa gatgaggcca 480gccgagtatt gggtagagga cgatgggtcc attgttaaga tcaggcacgg tgaaacattt 540gacgactaca tgaagttctt tgatggtctt cctgcctagg cccttttatc ttgttttggg 600caagcgtagc ccttttcatt tgatgagcgc atctcgtgga agattcgtgt gggaaaacta 660ttcacttgtt tgttatgtgg gtcatcccca tcaagcatgg gggtttttat ttgttagaat 720agagtccaac aagtttagtg attgtagaga ttgaatggac ttactgcatt gttatcaatt 780cttgtttata ctatataaag ggtccgactc ctcccaataa agttaaagaa tattgttgtt 840tacttttatc taaaaaaa 85856192PRTTriticum aestivum 56Phe Glu Leu Glu Tyr Leu Asn Ile Gly Gly Gly Leu Gly Ile Asp Tyr1 5 10 15His His Thr Gly Ala Val Leu Pro Thr Pro Met Asp Leu Ile Asn Thr20 25 30Val Arg Glu Leu Val Leu Ser Arg Asp Leu Thr Leu Ile Ile Glu Pro35 40 45Gly Arg Ser Leu Ile Ala Asn Thr Cys Cys Phe Val Asn Lys Val Thr50 55 60Gly Val Lys Ser Asn Gly Thr Lys Asn Phe Ile Val Val Asp Gly Ser65 70 75 80Met Ala Glu Leu Ile Arg Pro Ser Leu Tyr Gly Ala Tyr Gln His Ile85 90 95Glu Leu Val Ser Pro Ser Pro Gly Ala Glu Val Ala Thr Phe Asp Ile100 105 110Val Gly Pro Val Cys Glu Ser Ala Asp Phe Leu Gly Lys Asp Arg Glu115 120 125Leu Pro Thr Pro Asp Lys Gly Ala Gly Leu Val Val His Asp Ala Gly130 135 140Ala Tyr Cys Met Ser Met Ala Ser Thr Tyr Asn Leu Lys Met Arg Pro145 150 155 160Ala Glu Tyr Trp Val Glu Asp Asp Gly Ser Ile Val Lys Ile Arg His165 170 175Gly Glu Thr Phe Asp Asp Tyr Met Lys Phe Phe Asp Gly Leu Pro Ala180 185 19057526PRTArabidopsis thaliana 57Met Gly Gln Thr Asn Ser Glu Thr Gln Gln Ala Arg Leu Tyr Thr Gln1 5 10 15Asn Ser Gln Lys Gln Leu Leu Arg Ser Phe Leu Leu Leu His Leu Ile20 25 30Phe Gly Tyr Gln Ser His Lys Thr Leu Arg Met Ala Ala Ala Thr Gln35 40 45Phe Leu Ser Gln Pro Ser Ser Leu Asn Pro His Gln Leu Lys Asn Gln50 55 60Thr Ser Gln Arg Ser Arg Ser Ile Pro Val Leu Ser Leu Lys Ser Thr65 70 75 80Leu Lys Pro Leu Lys Arg Leu Ser Val Lys Ala Ala Val Val Ser Gln85 90 95Asn Ser Ser Lys Thr Val Thr Lys Phe Asp His Cys Phe Lys Lys Ser100 105 110Ser Asp Gly Phe Leu Tyr Cys Glu Gly Thr Lys Val Glu Asp Ile Met115 120 125Glu Ser Val Glu Arg Arg Pro Phe Tyr Leu Tyr Ser Lys Pro Gln Ile130 135 140Thr Arg Asn Leu Glu Ala Tyr Lys Glu Ala Leu Glu Gly Val Ser Ser145 150 155 160Val Ile Gly Tyr Ala Ile Lys Ala Asn Asn Asn Leu Lys Ile Leu Glu165 170 175His Leu Arg Ser Leu Gly Cys Gly Ala Val Leu Val Ser Gly Asn Glu180 185 190Leu Arg Leu Ala Leu Arg Ala Gly Phe Asp Pro Thr Lys Cys Ile Phe195 200 205Asn Gly Asn Gly Lys Ser Leu Glu Asp Leu Val Leu Ala Ala Gln Glu210 215 220Gly Val Phe Val Asn Val Asp Ser Glu Phe Asp Leu Asn Asn Ile Val225 230 235 240Glu Ala Ser Arg Ile Ser Gly Lys Gln Val Asn Val Leu Leu Arg Ile245 250 255Asn Pro Asp Val Asp Pro Gln Val His Pro Tyr Val Ala Thr Gly Asn260 265 270Lys Asn Ser Lys Phe Gly Ile Arg Asn Glu Lys Leu Gln Trp Phe Leu275 280 285Asp Gln Val Lys Ala His Pro Lys Glu Leu Lys Leu Val Gly Ala His290 295 300Cys His Leu Gly Ser Thr Ile Thr Lys Val Asp Ile Phe Arg Asp Ala305 310 315 320Ala Val Leu Met Ile Glu Tyr Ile Asp Glu Ile Arg Arg Gln Gly Phe325 330 335Glu Val Ser Tyr Leu Asn Ile Gly Gly Gly Leu Gly Ile Asp Tyr Tyr340 345 350His Ala Gly Ala Val Leu Pro Thr Pro Met Asp Leu Ile Asn Thr Val355 360 365Arg Glu Leu Val Leu Ser Arg Asp Leu Asn Leu Ile Ile Glu Pro Gly370 375 380Arg Ser Leu Ile Ala Asn Thr Cys Cys Phe Val Asn His Val Thr Gly385 390 395 400Val Lys Thr Asn Gly Thr Lys Asn Phe Ile Val Ile Asp Gly Ser Met405 410 415Ala Glu Leu Ile Arg Pro Ser Leu Tyr Asp Ala Tyr Gln His Ile Glu420 425 430Leu Val Ser Pro Pro Pro Ala Glu Ala Glu Val Thr Lys Phe Asp Val435 440 445Val Gly Pro Val Cys Glu Ser Ala Asp Phe Leu Gly Lys Asp Arg Glu450 455 460Leu Pro Thr Pro Pro Gln Gly Ala Gly Leu Val Val His Asp Ala Gly465 470 475 480Ala Tyr Cys Met Ser Met Ala Ser Thr Tyr Asn Leu Lys Met Arg Pro485 490 495Pro Glu Tyr Trp Val Glu Glu Asp Gly Ser Ile Thr Lys Ile Arg His500 505 510Ala Glu Thr Phe Asp Asp His Leu Arg Phe Phe Glu Gly Leu515 520 525581143DNAOryza sativa 58gcacgaggtc gccgccatcg ctgcccttcg cgccctcgat gtcaagtccc acgccgtctc 60catccacctc accaagggcc tccccctcgg ctccggcctc ggctcctccg ccgcctccgc 120cgccgccgct gccaaggccg ttgacgccct cttcggctcc ctcctacacc aagatgacct 180cgtcctcgcg ggcctcgagt ccgagaaagc cgtcagtggc ttccacgccg acaacatcgc 240cccggccatc ctcggcggct tcgtcctcgt ccgcagctac gaccccttcc acctcatccc 300gctctcctcc ccacctgccc tccgcctcca cttcgtcctc gtcacgcccg acttcgaggc 360gcccaccagc aagatgcgtg ccgcgctgcc caaacaggtg gccgtccacc agcacgtccg 420caactccagc caagcggccg cgcttgtcgc cgctgtgctg caaggggacg ccaccctcat 480cggctccgca atgtcctccg acggcatcgt ggagccaacc agggcgccgc tgattcctgg 540catggctgcg gtcaaggccg cggcgttgga agctggggca ttgggctgca ccatcagtgg 600agcagggcca actgctgtgg ctgtcattga cggggaggag aagggcgagg aggttggccg 660gaggatggtg gaggcattcg ccaatgccgg caatctcaaa gcaacagcta ctgttgctca 720gctcgataga gttggtgcca gggttatctc tacctccact ttggagtagg aagatctggg 780aggactgctc cggtaggtca aatttggaat ggctcacatg gacactagtg ggaggagaag 840aaggggggat tggtgtgttt tgtaattcct gggctgacca gaacgattgt cagtcagttg 900ggttgtgaat tgtgtgatgt agtagcaaac tgattcgtgc cggcaattga attgcaataa 960gctagtggtt gcagcatcac ctggcgaggc gtagctagga gatgcagaaa cagcattttg 1020acatgtgtgg gtgttgacat gcaacgaata aaatgaatga agctgaattg gggtttaaaa 1080aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaata 1140aaa 114359255PRTOryza sativa 59His Glu Val Ala Ala Ile Ala Ala Leu Arg Ala Leu Asp Val Lys Ser1 5 10 15His Ala Val Ser Ile His Leu Thr Lys Gly Leu Pro Leu Gly Ser Gly20 25 30Leu Gly Ser Ser Ala Ala Ser Ala Ala Ala Ala Ala Lys Ala Val Asp35 40 45Ala Leu Phe Gly Ser Leu Leu His Gln Asp Asp Leu Val Leu Ala Gly50 55 60Leu Glu Ser Glu Lys Ala Val Ser Gly Phe His Ala Asp Asn Ile Ala65 70 75 80Pro Ala Ile Leu Gly Gly Phe Val Leu Val Arg Ser Tyr Asp Pro Phe85 90 95His Leu Ile Pro Leu Ser Ser Pro Pro Ala Leu Arg Leu His Phe Val100 105 110Leu Val Thr Pro Asp Phe Glu Ala Pro Thr Ser Lys Met Arg Ala Ala115 120 125Leu Pro Lys Gln Val Ala Val His Gln His Val Arg Asn Ser Ser Gln130 135 140Ala Ala Ala Leu Val Ala Ala Val Leu Gln Gly Asp Ala Thr Leu Ile145 150 155 160Gly Ser Ala Met Ser Ser Asp Gly Ile Val Glu Pro Thr Arg Ala Pro165 170 175Leu Ile Pro Gly Met Ala Ala Val Lys Ala Ala Ala Leu Glu Ala Gly180 185 190Ala Leu Gly Cys Thr Ile Ser Gly Ala Gly Pro Thr Ala Val Ala Val195 200 205Ile Asp Gly Glu Glu Lys Gly Glu Glu Val Gly Arg Arg Met Val Glu210 215 220Ala Phe Ala Asn Ala Gly Asn Leu Lys Ala Thr Ala Thr Val Ala Gln225 230 235 240Leu Asp Arg Val Gly Ala Arg Val Ile Ser Thr Ser Thr Leu Glu245 250 25560370PRTArabidopsis thaliana 60Met Ala Ser Leu Cys Phe Gln Ser Pro Ser Lys Pro Ile Ser Tyr Phe1 5 10 15Gln Pro Lys Ser Asn Pro Ser Pro Pro Leu Phe Ala Lys Val Ser Val20 25 30Phe Arg Cys Arg Ala Ser Val Gln Thr Leu Val Ala Val Glu Pro Glu35 40 45Pro Val Phe Val Ser Val Lys Thr Phe Ala Pro Ala Thr Val Ala Asn50 55 60Leu Gly Pro Gly Phe Asp Phe Leu Gly Cys Ala Val Asp Gly Leu Gly65 70 75 80Asp His Val Thr Leu Arg Val Asp Pro Ser Val Arg Ala Gly Glu Val85 90 95Ser Ile Ser Glu Ile Thr Gly Thr Thr Thr Lys Leu Ser Thr Asn Pro100 105 110Leu Arg Asn Cys Ala Gly Ile Ala Ala Ile Ala Thr Met Lys Met Leu115 120 125Gly Ile Arg Ser Val Gly Leu Ser Leu Asp Leu His Lys Gly Leu Pro130 135 140Leu Gly Ser Gly Leu Gly Ser Ser Ala Ala Ser Ala Ala Ala Ala Ala145 150 155 160Val Ala Val Asn Glu Ile Phe Gly Arg Lys Leu Gly Ser Asp Gln Leu165 170 175Val Leu Ala Gly Leu Glu Ser Glu Ala Lys Val Ser Gly Tyr His Ala180 185 190Asp Asn Ile Ala Pro Ala Ile Met Gly Gly Phe Val Leu Ile Arg Asn195 200 205Tyr Glu Pro Leu Asp Leu Lys Pro Leu Lys Phe Pro Ser Asp Lys Asp210 215 220Leu Phe Phe Val Leu Val Ser Pro Glu Phe Glu Ala Pro Thr Lys Lys225 230 235 240Met Arg Ala Ala Leu Pro Thr Glu Ile Pro Met Val His His Val Trp245 250 255Asn Ser Ser Gln Ala Ala Ala Leu Val Ala Ala Val Leu Glu Gly Asp260 265 270Ala Val Met Leu Gly Lys Ala Leu Ser Ser Asp Lys Ile Val Glu Pro275 280 285Thr Arg Ala Pro Leu Ile Pro Gly Met Glu Ala Val Lys Lys Ala Ala290 295 300Leu Glu Ala Gly Ala Phe Gly Cys Thr Ile Ser Gly Ala Gly Pro Thr305 310 315 320Ala Val Ala Val Ile Asp Ser Glu Glu Lys Gly Gln Val Ile Gly Glu325 330 335Lys Met Val Glu Ala Phe Trp Lys Val Gly His Leu Lys Ser Val Ala340 345 350Ser Val Lys Lys Leu Asp Lys Val Gly Ala Arg Leu Val Asn Ser Val355 360 365Ser Arg370611508DNAZea mays 61aaggatggcg tcgtggtcgt cgccctcagc cgccgccaac gccgcctcgg gcgcccgatt 60cggccccttc ccgagcggag ggcagcggct cgcgccgtgt ccgtcgctcg tccgcggaac 120tcccgccccg acgctcgtcc tcaggctcca cccggacggc cgtggccatg gcctcctcgc 180gcacaccggc ccctctccct cctcgcggtg ccgcgccgtc gccgccgagg tcgggggcct 240caacatcgcc aacgacgtca cccagctcat cggcaacaca ccaatggtgt atctcaacaa 300cgtcgtcaag ggctctgtcg ccaatgtcgc tgctaagctc gagattatgg agccctgctg 360tagcgtcaag gacaggatag ggtacagtat gataaatgat gctgaacaga agggcttgat 420tactcctgga aagagtgttt tggtggaagc aacaagtgga aacacaggca ttggtcttgc 480tttcattgct gcttccaaag gatataagct gatactaaca atgccttcat caatgagcat 540ggagagaaga gtcctcctta gagcttttgg tgccgaactt gtccttactg atgctgcaaa 600agggatgaaa ggggccttag ataaggctac agagatttta aacaagacac caaattctta 660catgcttcaa cagttcgata accctgccaa ccctcaggta cattatgaga ctactggtcc 720agagatctgg gaggattcaa aggggaaggt ggatatattc attggtggaa ttggaacagg 780ggggacaata tctggtgccg gccgttttct caaggagaaa aatcctggaa ttaaggttat 840tggtattgag ccttctgaaa gtaacatact ctccggtgga aaacctggtc cacataagat 900ccagggaatc ggcgcaggat ttgttccaag gaacttggat agcgatattc ttgatgaagt 960aattgagata tcaagtgatg aagctgttga gacagcaaaa cagttggctg ttcaggaagg 1020attactggtt ggaatctcct ctggagcagc cgccgctgct gccataaagg ttgccaaaag 1080accagagaat gctggaaagc tgatagtggt tgtgtttccg agcttcggcg agaggtacct 1140ttcatctgtc ctctatcagt ccataagaga agaatgtgag aacatgcaac ctgagccatg 1200agggagccgt cactttaagc gggcatagta aatgtttctg aaataagacg cgtagccagc 1260atcagtttgc tccacttgga atcatttggc catgctcact ctatcctttc gctagcctct 1320atgaccggac ctaaactggt gtgtgagaaa catccacgac tgtcctccca actgctttcc 1380taaagccaaa cgataacact ctcaataatt gtctatacga ttgaagctga tttgattggt 1440aattgtaaac agcttgtctt tggatctttg aagtcaaaca aagtcagttg gttgaatcaa 1500aaaaaaaa 150862398PRTZea mays 62Met Ala Ser Trp Ser Ser Pro Ser Ala Ala Ala Asn Ala Ala Ser Gly1 5 10 15Ala Arg Phe Gly Pro Phe Pro Ser Gly Gly Gln Arg Leu Ala Pro Cys20 25 30Pro Ser Leu Val Arg Gly Thr Pro Ala Pro Thr Leu Val Leu Arg Leu35 40 45His Pro Asp Gly Arg Gly His Gly Leu Leu Ala His Thr Gly Pro Ser50 55 60Pro Ser Ser Arg Cys Arg Ala Val Ala Ala Glu Val Gly Gly Leu Asn65 70 75 80Ile Ala Asn Asp Val Thr Gln Leu Ile Gly Asn Thr Pro Met Val Tyr85 90 95Leu Asn Asn Val Val Lys Gly Ser Val Ala Asn Val Ala Ala Lys Leu100 105 110Glu Ile Met Glu Pro Cys Cys Ser Val Lys Asp Arg Ile Gly Tyr Ser115 120 125Met Ile Asn Asp Ala Glu Gln Lys Gly Leu Ile Thr Pro Gly Lys Ser130 135 140Val Leu Val Glu Ala Thr Ser Gly Asn Thr Gly Ile Gly Leu Ala Phe145 150 155 160Ile Ala Ala Ser Lys Gly Tyr Lys Leu Ile Leu Thr Met Pro Ser Ser165 170 175Met Ser Met Glu Arg Arg Val Leu Leu Arg Ala Phe Gly Ala Glu Leu180 185 190Val Leu Thr Asp Ala Ala Lys Gly Met Lys Gly Ala Leu Asp Lys Ala195 200 205Thr Glu Ile Leu Asn Lys Thr Pro Asn Ser Tyr Met Leu Gln Gln Phe210 215 220Asp Asn Pro Ala Asn Pro Gln Val His Tyr Glu Thr Thr Gly Pro Glu225 230 235 240Ile Trp Glu Asp Ser Lys Gly Lys Val Asp Ile Phe Ile Gly Gly Ile245 250 255Gly Thr Gly Gly Thr Ile Ser Gly Ala Gly Arg Phe Leu Lys Glu Lys260 265 270Asn Pro Gly Ile Lys Val Ile Gly Ile Glu Pro Ser Glu Ser Asn Ile275 280 285Leu Ser Gly Gly Lys Pro Gly Pro His Lys Ile Gln Gly Ile Gly Ala290 295 300Gly Phe Val Pro Arg Asn Leu Asp Ser Asp Ile Leu Asp Glu Val Ile305 310 315 320Glu Ile Ser Ser Asp Glu Ala Val Glu Thr Ala Lys Gln Leu Ala Val325 330 335Gln Glu Gly Leu Leu Val Gly Ile Ser Ser Gly Ala Ala Ala Ala Ala340 345 350Ala Ile Lys Val Ala Lys Arg Pro Glu Asn Ala Gly Lys Leu Ile Val355 360 365Val Val Phe Pro Ser Phe Gly Glu Arg Tyr Leu Ser Ser Val Leu Tyr370 375 380Gln Ser Ile Arg Glu Glu Cys Glu Asn Met Gln Pro Glu Pro385 390 395631522DNAOryza sativa 63gcacgaggtt ctaactacgg aactactccc ctatccaaca cctccgagtc cgagcaacgc 60aagatggcgt cgtggtcgtc gcccgtcgcc gccgccgcct tgcaggtcca tttcgggtcc 120tcctgcttct tctccgcccg atcgccacga cagaccctcc tcctaccacc tctcgcccgc 180aaccctacac tgaccatcca gccccggccc catcccttcc ggaacatcaa ctcctcctcc 240tcctccagct ggatgtgcca cgccgtcgcc gccgaggtcg agggcctcaa catcgccgac 300gacgtcaccc agctcatcgg caagactcca atggtatatc tcaacaacat cgtcaaggga 360tgtgttgcca atgtcgctgc taagctcgag attatggagc cctgttgcag tgtcaaggac 420aggataggat acagtatgat ttctgatgcg gaagagaaag gcttgataac tcctggaaag 480agtgttttgg tggaaccaac aagtggaaat acaggcattg gtcttgcctt cattgctgct 540tccagaggat ataaattaat attgaccatg cctgcatcaa tgagcatgga gagaagagtt 600ctactcaaag cttttggcgc tgaacttgtc cttactgatg ccgcaaaagg gatgaagggg 660gctgtagata aggctacaga gattttaaat aagacacctg atgcctatat gctgcagcag 720tttgacaacc ctgccaaccc aaaggtacat tatgagacta ctgggccaga aatctgggag 780gattctaaag ggaaggtgga tgtattcatt ggtggaattg gaacaggtgg aacaatatct 840ggtgctggcc gtttcctgaa agagaaaaat cctggaatta aggttattgg tattgagcct 900tctgagagta acatactctc tggtggaaaa cctggcccac ataagattca aggcattggg 960gcaggatttg ttccaaggaa cttggatagt gaagttctcg atgaagtgat tgagatatct 1020agtgatgagg ctgttgagac agcaaagcaa ttggctcttc aggaaggatt actggttgga 1080atttcatctg gggcagcagc agcagctgcc attaaagttg caaaaagacc agaaaatgct 1140ggaaagttgg tagtggttgt gtttccaagc tttggtgaga ggtacctttc atctatcctt 1200tttcagtcga taagagaaga atgtgagaag ttgcaacctg aaccatgagc ctaacttcag 1260tgttcacaac atcataattg tttctgagat ttctggccat tagttttttt ttctgagaag 1320tatcatacca ctccatagct gtttgttcga taaataaaac agttaccttt gcacttataa 1380tgaggcttgt gagggtactg tgaaatttct ctgaacatct tctactcttc tcttttatcc 1440ttaaatcaat ctgggagcag tttgtaatac atacgtaaat ttaaagctgg gtgtttggta 1500attgtaaaaa aaaaaaaaaa aa 152264415PRTOryza sativa 64Ala Arg Gly Ser Asn Tyr Gly Thr Thr Pro Leu Ser Asn Thr Ser Glu1 5 10 15Ser Glu Gln Arg Lys Met Ala Ser Trp Ser Ser Pro Val Ala Ala Ala20 25 30Ala Leu Gln Val His Phe Gly

Ser Ser Cys Phe Phe Ser Ala Arg Ser35 40 45Pro Arg Gln Thr Leu Leu Leu Pro Pro Leu Ala Arg Asn Pro Thr Leu50 55 60Thr Ile Gln Pro Arg Pro His Pro Phe Arg Asn Ile Asn Ser Ser Ser65 70 75 80Ser Ser Ser Trp Met Cys His Ala Val Ala Ala Glu Val Glu Gly Leu85 90 95Asn Ile Ala Asp Asp Val Thr Gln Leu Ile Gly Lys Thr Pro Met Val100 105 110Tyr Leu Asn Asn Ile Val Lys Gly Cys Val Ala Asn Val Ala Ala Lys115 120 125Leu Glu Ile Met Glu Pro Cys Cys Ser Val Lys Asp Arg Ile Gly Tyr130 135 140Ser Met Ile Ser Asp Ala Glu Glu Lys Gly Leu Ile Thr Pro Gly Lys145 150 155 160Ser Val Leu Val Glu Pro Thr Ser Gly Asn Thr Gly Ile Gly Leu Ala165 170 175Phe Ile Ala Ala Ser Arg Gly Tyr Lys Leu Ile Leu Thr Met Pro Ala180 185 190Ser Met Ser Met Glu Arg Arg Val Leu Leu Lys Ala Phe Gly Ala Glu195 200 205Leu Val Leu Thr Asp Ala Ala Lys Gly Met Lys Gly Ala Val Asp Lys210 215 220Ala Thr Glu Ile Leu Asn Lys Thr Pro Asp Ala Tyr Met Leu Gln Gln225 230 235 240Phe Asp Asn Pro Ala Asn Pro Lys Val His Tyr Glu Thr Thr Gly Pro245 250 255Glu Ile Trp Glu Asp Ser Lys Gly Lys Val Asp Val Phe Ile Gly Gly260 265 270Ile Gly Thr Gly Gly Thr Ile Ser Gly Ala Gly Arg Phe Leu Lys Glu275 280 285Lys Asn Pro Gly Ile Lys Val Ile Gly Ile Glu Pro Ser Glu Ser Asn290 295 300Ile Leu Ser Gly Gly Lys Pro Gly Pro His Lys Ile Gln Gly Ile Gly305 310 315 320Ala Gly Phe Val Pro Arg Asn Leu Asp Ser Glu Val Leu Asp Glu Val325 330 335Ile Glu Ile Ser Ser Asp Glu Ala Val Glu Thr Ala Lys Gln Leu Ala340 345 350Leu Gln Glu Gly Leu Leu Val Gly Ile Ser Ser Gly Ala Ala Ala Ala355 360 365Ala Ala Ile Lys Val Ala Lys Arg Pro Glu Asn Ala Gly Lys Leu Val370 375 380Val Val Val Phe Pro Ser Phe Gly Glu Arg Tyr Leu Ser Ser Ile Leu385 390 395 400Phe Gln Ser Ile Arg Glu Glu Cys Glu Lys Leu Gln Pro Glu Pro405 410 41565383PRTSpinacia oleracea 65Met Ala Ser Leu Val Asn Asn Ala Tyr Ala Ala Ile Arg Thr Ser Lys1 5 10 15Leu Glu Leu Arg Glu Val Lys Asn Leu Ala Asn Phe Arg Val Gly Pro20 25 30Pro Ser Ser Leu Ser Cys Asn Asn Phe Lys Lys Val Ser Ser Ser Pro35 40 45Ile Thr Cys Lys Ala Val Ser Leu Ser Pro Pro Ser Thr Ile Glu Gly50 55 60Leu Asn Ile Ala Glu Asp Val Ser Gln Leu Ile Gly Lys Thr Pro Met65 70 75 80Val Tyr Leu Asn Asn Val Ser Lys Gly Ser Val Ala Asn Ile Ala Ala85 90 95Lys Leu Glu Ser Met Glu Pro Cys Cys Ser Val Lys Asp Arg Ile Gly100 105 110Tyr Ser Met Ile Asp Asp Ala Glu Gln Lys Gly Val Ile Thr Pro Gly115 120 125Lys Thr Thr Leu Val Glu Pro Thr Ser Gly Asn Thr Gly Ile Gly Leu130 135 140Ala Phe Ile Ala Ala Ala Arg Gly Tyr Lys Ile Thr Leu Thr Met Pro145 150 155 160Ala Ser Met Ser Met Glu Arg Arg Val Ile Leu Lys Ala Phe Gly Ala165 170 175Glu Leu Val Leu Thr Asp Pro Ala Lys Gly Met Lys Gly Ala Val Glu180 185 190Lys Ala Glu Glu Ile Leu Lys Lys Thr Pro Asp Ser Tyr Met Leu Gln195 200 205Gln Phe Asp Asn Pro Ala Asn Pro Lys Ile His Tyr Glu Thr Thr Gly210 215 220Pro Glu Ile Trp Glu Asp Thr Lys Gly Lys Val Asp Ile Phe Val Ala225 230 235 240Gly Ile Gly Thr Gly Gly Thr Ile Ser Gly Val Gly Arg Tyr Leu Lys245 250 255Glu Arg Asn Pro Gly Val Gln Val Ile Gly Ile Glu Pro Thr Glu Ser260 265 270Asn Ile Leu Ser Gly Gly Lys Pro Gly Pro His Lys Ile Gln Gly Leu275 280 285Gly Ala Gly Phe Val Pro Ser Asn Leu Asp Leu Gly Val Met Asp Glu290 295 300Val Ile Glu Val Ser Ser Glu Glu Ala Val Glu Met Ala Lys Gln Leu305 310 315 320Ala Met Lys Glu Gly Leu Leu Val Gly Ile Ser Ser Gly Ala Ala Ala325 330 335Ala Ala Ala Val Arg Ile Gly Lys Arg Pro Glu Asn Ala Gly Lys Leu340 345 350Ile Ala Val Val Phe Pro Ser Phe Gly Glu Arg Tyr Leu Ser Ser Ile355 360 365Leu Phe Gln Ser Ile Arg Glu Glu Cys Glu Asn Met Lys Pro Glu370 375 38066386PRTSolanum tuberosum 66Met Ala Ser Phe Ile Asn Asn Pro Leu Thr Ser Leu Cys Asn Thr Lys1 5 10 15Ser Glu Arg Asn Asn Leu Phe Lys Ile Ser Leu Tyr Glu Ala Gln Ser20 25 30Leu Gly Phe Ser Lys Leu Asn Gly Ser Arg Lys Val Ala Phe Pro Ser35 40 45Val Val Cys Lys Ala Val Ser Val Pro Thr Lys Ser Ser Thr Glu Ile50 55 60Glu Gly Leu Asn Ile Ala Glu Asp Val Thr Gln Leu Ile Gly Asn Thr65 70 75 80Pro Met Val Tyr Leu Asn Thr Ile Ala Lys Gly Cys Val Ala Asn Ile85 90 95Ala Ala Lys Leu Glu Ile Met Glu Pro Cys Cys Ser Val Lys Asp Arg100 105 110Ile Gly Phe Ser Met Ile Val Asp Ala Glu Glu Lys Gly Leu Ile Ser115 120 125Pro Gly Lys Thr Val Leu Val Glu Pro Thr Ser Gly Asn Thr Gly Ile130 135 140Gly Leu Ala Phe Ile Ala Ala Ser Arg Gly Tyr Lys Leu Ile Leu Thr145 150 155 160Met Pro Ala Ser Met Ser Leu Glu Arg Arg Val Ile Leu Lys Ala Phe165 170 175Gly Ala Glu Leu Val Leu Thr Asp Pro Ala Lys Gly Met Lys Gly Ala180 185 190Val Ser Lys Ala Glu Glu Ile Leu Asn Asn Thr Pro Asp Ala Tyr Ile195 200 205Leu Gln Gln Phe Asp Asn Pro Ala Asn Pro Lys Ile His Tyr Glu Thr210 215 220Thr Gly Pro Glu Ile Trp Glu Asp Thr Lys Gly Lys Ile Asp Ile Leu225 230 235 240Val Ala Gly Ile Gly Thr Gly Gly Thr Ile Thr Gly Thr Gly Arg Phe245 250 255Leu Lys Glu Gln Asn Pro Asn Ile Lys Ile Ile Gly Val Glu Pro Thr260 265 270Glu Ser Asn Val Leu Ser Gly Gly Lys Pro Gly Pro His Lys Ile Gln275 280 285Gly Ile Gly Ala Gly Phe Ile Pro Gly Asn Leu Asp Gln Asp Val Met290 295 300Asp Glu Val Ile Glu Ile Ser Ser Asp Glu Ala Val Glu Thr Ala Arg305 310 315 320Thr Leu Ala Leu Gln Glu Gly Leu Leu Val Gly Ile Ser Ser Gly Ala325 330 335Ala Ala Leu Ala Ala Ile Gln Val Gly Lys Arg Pro Glu Asn Ala Gly340 345 350Lys Leu Ile Gly Val Val Phe Pro Ser Tyr Gly Glu Arg Tyr Leu Ser355 360 365Ser Ile Leu Phe Gln Ser Ile Arg Glu Glu Cys Glu Lys Met Lys Pro370 375 380Glu Leu385671581DNAZea mays 67ggccgtggct tactggcttc cacccacagc cttcgcactt ccctccttcc tcgcaaatgg 60ccgtcgccgt ccccaacgct cccggccgcc tcttccttct ccaatccacc ccgttcccga 120accctagcag ctcggcatcc gccgctcgag cccaatcctt ccgcgtacca cccctccgcc 180tctcgctatt ccgacgcatg gctgggcgct cgctgacggt gatcgcaggc gcctccggcg 240gctccgaacg agatctcagc gcctccgcag tctccgtgga ggccctggac tccgtcgcct 300ccgattctga cttagagacg aaggagccca gtgtgtcgac gatgctgacg agcttcgaga 360actcgttcga caagtatggg gctctgagca caccgctgta ccagaccgcc acctttaagc 420agccttcagc tacagattat ggaacttatg attacactag aagtggtaac cctactcgtg 480atgttctcca gagcctcatg gctaagcttg agaaagcaga tcaagcattc tgcttcacca 540gcgggatggc ggcgttagct gcagtaaaac acctccttca ggctggacaa gaaatagttg 600ctggtgagga catatatggt ggttctgatc gtctactctc gcaagttgtg ccaagaaatg 660gaatagttgt aaaacgagta gatacaacga aaattagtga tgtggtgtct gcaattggac 720cctccactag actggtttgg ctcgaaagtc ccacgaaccc tcgtcagcaa attactgaca 780ttaagacaat ctcagagata gcgcattctc atggtgctct tgttttggtt gacaacagca 840tcatgtctcc agtgctctcc cgtcctatag aactgggagc tgatatcgtg atgcactcgg 900ctaccaaatt tatagcggga catagtgatc ttatggctgg aattcttgca gtgaagggtg 960agagtttggc taaagaggta gggtttctgc aaaatgctga agggtcgggt ctggcacctt 1020ttgactgctg gctttgcttg aggggaatca aaaccatggc tctgcgggtg gagaaacaac 1080aggctaatgc ccagaagatt gctgaattcc tggcgtctca cccgagggtc aagcaagtaa 1140actacgctgg gcttcctgac catcctgggc gagctttaca ctattcccag gcaaagggag 1200cgggctctgt tctcagtttt ctcaccggct cactggccct ctcaaagcac gtcgtggaga 1260ccaccaagta cttcagcgta acagtcagct tcgggagcgt gaagtccctc atcagcctgc 1320cgtgcttcat gtcccacgca tcaatccctg cctcggtccg cgaggagcgt ggcctaaccg 1380acgacctcgt ccggatatcg gtcggcatcg aggatgtcga ggacctcatc gccgatctgg 1440accgcgcgct cagaactggc ccggtgtaga catcgccgat ccttaggtca tgtcaagcta 1500tcttttgatg attcattggt tgactgcttg cgtgatgata ataatgggaa tgttgcttgg 1560ataaaaaaaa aaaaaaaaaa a 158168470PRTZea mays 68Met Ala Val Ala Val Pro Asn Ala Pro Gly Arg Leu Phe Leu Leu Gln1 5 10 15Ser Thr Pro Phe Pro Asn Pro Ser Ser Ser Ala Ser Ala Ala Arg Ala20 25 30Gln Ser Phe Arg Val Pro Pro Leu Arg Leu Ser Leu Phe Arg Arg Met35 40 45Ala Gly Arg Ser Leu Thr Val Ile Ala Gly Ala Ser Gly Gly Ser Glu50 55 60Arg Asp Leu Ser Ala Ser Ala Val Ser Val Glu Ala Leu Asp Ser Val65 70 75 80Ala Ser Asp Ser Asp Leu Glu Thr Lys Glu Pro Ser Val Ser Thr Met85 90 95Leu Thr Ser Phe Glu Asn Ser Phe Asp Lys Tyr Gly Ala Leu Ser Thr100 105 110Pro Leu Tyr Gln Thr Ala Thr Phe Lys Gln Pro Ser Ala Thr Asp Tyr115 120 125Gly Thr Tyr Asp Tyr Thr Arg Ser Gly Asn Pro Thr Arg Asp Val Leu130 135 140Gln Ser Leu Met Ala Lys Leu Glu Lys Ala Asp Gln Ala Phe Cys Phe145 150 155 160Thr Ser Gly Met Ala Ala Leu Ala Ala Val Lys His Leu Leu Gln Ala165 170 175Gly Gln Glu Ile Val Ala Gly Glu Asp Ile Tyr Gly Gly Ser Asp Arg180 185 190Leu Leu Ser Gln Val Val Pro Arg Asn Gly Ile Val Val Lys Arg Val195 200 205Asp Thr Thr Lys Ile Ser Asp Val Val Ser Ala Ile Gly Pro Ser Thr210 215 220Arg Leu Val Trp Leu Glu Ser Pro Thr Asn Pro Arg Gln Gln Ile Thr225 230 235 240Asp Ile Lys Thr Ile Ser Glu Ile Ala His Ser His Gly Ala Leu Val245 250 255Leu Val Asp Asn Ser Ile Met Ser Pro Val Leu Ser Arg Pro Ile Glu260 265 270Leu Gly Ala Asp Ile Val Met His Ser Ala Thr Lys Phe Ile Ala Gly275 280 285His Ser Asp Leu Met Ala Gly Ile Leu Ala Val Lys Gly Glu Ser Leu290 295 300Ala Lys Glu Val Gly Phe Leu Gln Asn Ala Glu Gly Ser Gly Leu Ala305 310 315 320Pro Phe Asp Cys Trp Leu Cys Leu Arg Gly Ile Lys Thr Met Ala Leu325 330 335Arg Val Glu Lys Gln Gln Ala Asn Ala Gln Lys Ile Ala Glu Phe Leu340 345 350Ala Ser His Pro Arg Val Lys Gln Val Asn Tyr Ala Gly Leu Pro Asp355 360 365His Pro Gly Arg Ala Leu His Tyr Ser Gln Ala Lys Gly Ala Gly Ser370 375 380Val Leu Ser Phe Leu Thr Gly Ser Leu Ala Leu Ser Lys His Val Val385 390 395 400Glu Thr Thr Lys Tyr Phe Ser Val Thr Val Ser Phe Gly Ser Val Lys405 410 415Ser Leu Ile Ser Leu Pro Cys Phe Met Ser His Ala Ser Ile Pro Ala420 425 430Ser Val Arg Glu Glu Arg Gly Leu Thr Asp Asp Leu Val Arg Ile Ser435 440 445Val Gly Ile Glu Asp Val Glu Asp Leu Ile Ala Asp Leu Asp Arg Ala450 455 460Leu Arg Thr Gly Pro Val465 470691685DNAOryza sativa 69aggcaaccat gagcgccgcc gccgccgccg ccgccgccgc cgcaatcccc acctctctcg 60gccgcctctt ccacctccgc cccaccccga acccctcccg gaaccttagc ggcagctcag 120cgcaacccct cctccgcctc agctaccacc cacgcctcac gctctctcgc cgcatggagg 180cgccggcggc gatcgccgac tcccacggcg gcggcgacct gagcgcgtcc gcggtcggcg 240cggaggcgct gggcgccgtc gccgctccgg atttcgatgt ggagatgaag gagcctagcg 300tggcgacgat actgacgagc ttcgagaact cgttcgatgg gttcgggtct atgagcacgc 360cgctgtacca gacggccacg tttaagcagc cttcagcaac cgataatgga ccttatgatt 420acactagaag tggtaaccct acacgtgatg ttctccaaag ccttatggct aagcttgaga 480aggcggatca ggcattctgc ttcaccagtg ggatggcagc actagctgca gtaacacacc 540tccttaagtc tggacaagaa atagttgctg gagaggacat atatggtggc tcagaccgtc 600tgctctcaca agttgccccg agacatggga ttgtagtaaa acgaattgat acaaccaaaa 660ttagtgaggt aacttctgca attgggccct tgactaaact agtatggctt gaaagtccca 720ccaatccccg tctacaaatt actgatataa agaaaatagc agagatagct cattaccatg 780gtgctcttgt tttagtagac aacagcatca tgtctcctgt gctctcccgt cctctagaac 840ttggagcaga tattgttatg cactcagcaa ccaaatttat agctggacat agcgatctta 900tggctggaat tcttgcggtg aagggtgaaa gcagcttggc taaagagatt gcatttctac 960aaaatgctga aggatcaggt ttggcaccat ttgattgctg gctttgtttg agaggaatca 1020aaaccatggc tttgcgggtg gagaagcagc aggctaatgc tcagaagatt gctgaatttc 1080tagcttctca tccaagagta aagaaagtga actatgcagg acttcctgat catcctggac 1140gatctctaca ctattcccag gcaaagggag cgggttcagt tctcagtttc ctaactggtt 1200cattagctct ctcaaaacat gttgttgaga ccacaaagta cttcaatgta acagttagct 1260ttggaagtgt gaaatcgctc attagcctgc catgcttcat gtcacacgcc agcatccctt 1320ctgcggttcg cgaggagcgc ggcctgacag acgatctagt caggatatcg gttggaattg 1380aggatgccga cgacctcata gcggatcttg atcatgctct ccggtctggt ccagcttaga 1440gcctgtgaat tctgtgccct tcctgttcgt tagggatgta gatgtggtca tgtgggtgct 1500atctgtgtgg gtgattgatt cattggtcaa ctcaataagc tgctgtgtca tcgagggaat 1560aaagacaatc tatcccaaat tttttaacac catatggtga ccaactgacc atgatatggt 1620cttaatcaat tgatatttat agaaggtttc tttgaactgc aaaaaaaaaa aaaaaaaaaa 1680aaaaa 168570476PRTOryza sativa 70Met Ser Ala Ala Ala Ala Ala Ala Ala Ala Ala Ala Ile Pro Thr Ser1 5 10 15Leu Gly Arg Leu Phe His Leu Arg Pro Thr Pro Asn Pro Ser Arg Asn20 25 30Leu Ser Gly Ser Ser Ala Gln Pro Leu Leu Arg Leu Ser Tyr His Pro35 40 45Arg Leu Thr Leu Ser Arg Arg Met Glu Ala Pro Ala Ala Ile Ala Asp50 55 60Ser His Gly Gly Gly Asp Leu Ser Ala Ser Ala Val Gly Ala Glu Ala65 70 75 80Leu Gly Ala Val Ala Ala Pro Asp Phe Asp Val Glu Met Lys Glu Pro85 90 95Ser Val Ala Thr Ile Leu Thr Ser Phe Glu Asn Ser Phe Asp Gly Phe100 105 110Gly Ser Met Ser Thr Pro Leu Tyr Gln Thr Ala Thr Phe Lys Gln Pro115 120 125Ser Ala Thr Asp Asn Gly Pro Tyr Asp Tyr Thr Arg Ser Gly Asn Pro130 135 140Thr Arg Asp Val Leu Gln Ser Leu Met Ala Lys Leu Glu Lys Ala Asp145 150 155 160Gln Ala Phe Cys Phe Thr Ser Gly Met Ala Ala Leu Ala Ala Val Thr165 170 175His Leu Leu Lys Ser Gly Gln Glu Ile Val Ala Gly Glu Asp Ile Tyr180 185 190Gly Gly Ser Asp Arg Leu Leu Ser Gln Val Ala Pro Arg His Gly Ile195 200 205Val Val Lys Arg Ile Asp Thr Thr Lys Ile Ser Glu Val Thr Ser Ala210 215 220Ile Gly Pro Leu Thr Lys Leu Val Trp Leu Glu Ser Pro Thr Asn Pro225 230 235 240Arg Leu Gln Ile Thr Asp Ile Lys Lys Ile Ala Glu Ile Ala His Tyr245 250 255His Gly Ala Leu Val Leu Val Asp Asn Ser Ile Met Ser Pro Val Leu260 265 270Ser Arg Pro Leu Glu Leu Gly Ala Asp Ile Val Met His Ser Ala Thr275 280 285Lys Phe Ile Ala Gly His Ser Asp Leu Met Ala Gly Ile Leu Ala Val290 295 300Lys Gly Glu Ser Ser Leu Ala Lys Glu Ile Ala Phe Leu Gln Asn Ala305 310 315 320Glu Gly Ser Gly Leu Ala Pro Phe Asp Cys Trp Leu Cys Leu Arg Gly325 330 335Ile Lys Thr Met Ala Leu Arg Val Glu Lys Gln Gln Ala Asn Ala Gln340 345 350Lys Ile Ala Glu Phe Leu Ala Ser His Pro Arg Val Lys Lys Val Asn355 360 365Tyr Ala Gly Leu Pro Asp His Pro Gly Arg Ser Leu His Tyr Ser Gln370 375 380Ala Lys Gly Ala Gly Ser Val Leu Ser Phe Leu Thr Gly Ser Leu Ala385 390 395 400Leu Ser Lys His Val Val Glu Thr Thr Lys Tyr Phe Asn Val Thr Val405 410 415Ser Phe Gly Ser Val Lys Ser Leu Ile Ser Leu Pro Cys Phe Met Ser420

425 430His Ala Ser Ile Pro Ser Ala Val Arg Glu Glu Arg Gly Leu Thr Asp435 440 445Asp Leu Val Arg Ile Ser Val Gly Ile Glu Asp Ala Asp Asp Leu Ile450 455 460Ala Asp Leu Asp His Ala Leu Arg Ser Gly Pro Ala465 470 475711699DNATriticum aestivum 71gcacgagagc gtggccacga tactgaccag cttcgagaac tcgttcgaca agtatggggc 60tctcagcacg ccgctgtacc agacggccac cttcaagcag ccttcagcaa ccgttaatgg 120agcttatgat tatactagaa gtggcaaccc tactcgtgat gttctccaga gccttatggc 180taagctcgag aaggcagacc aagcattctg cttcactagt gggatggcat cactggctgc 240agtaacacac ctccttcagg ctggacaaga aatagttgct ggagaggaca tatatggtgg 300ctctgatcgt ctgctctcac aagttgtccc aagaaatgga attgtagtaa aacgggtcga 360tacaactaaa attaacgacg tgactgctgc aatcggaccc ttgactagac tagtttggct 420tgaaagtccc accaatcctc gtcaacaaat tactgatata aagaaaatct cagagatagc 480tcattctcat ggtgcacttg ttttggtgga caacagtatc atgtctccag tgctatcctg 540gcctatagaa cttggagcag atattgtgat gcactcagct accaaattta tagctggaca 600cagtgatctt atggctggaa ttcttgctgt aaagggtgaa agcttggcta aggagattgc 660atttctacaa aacgctgaag gttctggttt ggcacctttt gattgttggc tttgcttgag 720agggatcaaa accatggcct tacgggtgga aaagcaacag gataatgccc agaagattgc 780tgaattctta gcttctcatc caagggtcaa gcaagtgaat tatgctggac ttcctgatca 840tcctggccga tctttacact actctcaggc aaagggagcg ggctctgtcc tcagtttcca 900aactggttca ttgtctctct caaagcatgt tgttgagaca accaagtact tcaacgtaac 960agttagcttc ggaagtgtga agtcactcat aagcttgccc tgcttcatgt cgcacgcgag 1020catcccttcc tcggtgcgag aggagcgtgg gttgactgat gatctagtac ggatatcggt 1080gggtattgag gatgtggatg acctcatagc tgatcttgat tacgcgctca ggtccggtcc 1140agcatagatc atacaaaatc tggactatgg cgcttcgggt tctagttaat caagttgtag 1200atgtgatatg cattggtgat tcatttgtta agctgcaaca gtaataataa acttctgcac 1260gagtattttc tgaaatgacg agcccacggt tgtatgtgtt gttcctcata ggcttcaaca 1320gaaaaaccct gaggccaact gacaagtagc aacattcata aacttcacaa catcgatact 1380tggttctgcc catgttcatt tttcttggct gccattgtga cggctttgta gctcaagtag 1440gaaggagtga catggccgtt ggttgatggg gagaaaagga gttggttcgt cggatcgatc 1500cgtgtaggcg cttgtgtatt ttgtatatgg tgtttttcgt ctgtgcaggt gagtctgtgt 1560atacatctgg agactggatt attcatggtc attggtgtgg cggtgaagaa taatgtgacg 1620attcttttgt agtgtatcta agaactgtga tgttcttgtg caaaaaaaaa aaaaaaaaaa 1680aaaaaaaaaa aaaaaaaaa 169972381PRTTriticum aestivum 72His Glu Ser Val Ala Thr Ile Leu Thr Ser Phe Glu Asn Ser Phe Asp1 5 10 15Lys Tyr Gly Ala Leu Ser Thr Pro Leu Tyr Gln Thr Ala Thr Phe Lys20 25 30Gln Pro Ser Ala Thr Val Asn Gly Ala Tyr Asp Tyr Thr Arg Ser Gly35 40 45Asn Pro Thr Arg Asp Val Leu Gln Ser Leu Met Ala Lys Leu Glu Lys50 55 60Ala Asp Gln Ala Phe Cys Phe Thr Ser Gly Met Ala Ser Leu Ala Ala65 70 75 80Val Thr His Leu Leu Gln Ala Gly Gln Glu Ile Val Ala Gly Glu Asp85 90 95Ile Tyr Gly Gly Ser Asp Arg Leu Leu Ser Gln Val Val Pro Arg Asn100 105 110Gly Ile Val Val Lys Arg Val Asp Thr Thr Lys Ile Asn Asp Val Thr115 120 125Ala Ala Ile Gly Pro Leu Thr Arg Leu Val Trp Leu Glu Ser Pro Thr130 135 140Asn Pro Arg Gln Gln Ile Thr Asp Ile Lys Lys Ile Ser Glu Ile Ala145 150 155 160His Ser His Gly Ala Leu Val Leu Val Asp Asn Ser Ile Met Ser Pro165 170 175Val Leu Ser Trp Pro Ile Glu Leu Gly Ala Asp Ile Val Met His Ser180 185 190Ala Thr Lys Phe Ile Ala Gly His Ser Asp Leu Met Ala Gly Ile Leu195 200 205Ala Val Lys Gly Glu Ser Leu Ala Lys Glu Ile Ala Phe Leu Gln Asn210 215 220Ala Glu Gly Ser Gly Leu Ala Pro Phe Asp Cys Trp Leu Cys Leu Arg225 230 235 240Gly Ile Lys Thr Met Ala Leu Arg Val Glu Lys Gln Gln Asp Asn Ala245 250 255Gln Lys Ile Ala Glu Phe Leu Ala Ser His Pro Arg Val Lys Gln Val260 265 270Asn Tyr Ala Gly Leu Pro Asp His Pro Gly Arg Ser Leu His Tyr Ser275 280 285Gln Ala Lys Gly Ala Gly Ser Val Leu Ser Phe Gln Thr Gly Ser Leu290 295 300Ser Leu Ser Lys His Val Val Glu Thr Thr Lys Tyr Phe Asn Val Thr305 310 315 320Val Ser Phe Gly Ser Val Lys Ser Leu Ile Ser Leu Pro Cys Phe Met325 330 335Ser His Ala Ser Ile Pro Ser Ser Val Arg Glu Glu Arg Gly Leu Thr340 345 350Asp Asp Leu Val Arg Ile Ser Val Gly Ile Glu Asp Val Asp Asp Leu355 360 365Ile Ala Asp Leu Asp Tyr Ala Leu Arg Ser Gly Pro Ala370 375 380

Claims

1. An isolated polynucleotide that encodes a plant cysteine γ synthase having amino acid sequence identity of at least 95% based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:31, 62, and 64.

2. The polynucleotide of claim 1 wherein the polynucleotide encodes a polypeptide selected from the group consisting of SEQ ID NOs: NOs:31, 62, and 64.

3. The polynucleotide of claim 1, wherein the polynucleotide comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:30, 61, and 63.

4. An isolated complement of the polynucleotide of claim 1, wherein (a) the complement and the polynucleotide consist of the same number of nucleotides, and (b) the nucleotide sequences of the complement and the polynucleotide have 100% complementarity.

5. An isolated nucleic acid molecule that (1) comprises at least 180 nucleotides (2) remains hybridized with a polynucleotide having a nucleotide sequence selected from the group consisting of SEQ ID NO:30, 61, and 63 under a wash condition of 0.1.times.SSC, 0.1% SDS, and 65.degree. C., and encodes a plant cysteine γ synthase.

6. A cell comprising the polynucleotide of claim 1.

7. The cell of claim 6, wherein the cell is selected from the group consisting of a yeast cell, a bacterial cell and a plant cell.

8. A transgenic plant comprising the polynucleotide of claim 1.

9. A method for transforming a cell comprising introducing into a cell the polynucleotide of claim 1.

10. A method for producing a transgenic plant comprising (a) transforming a plant cell with the polynucleotide of claim 1, and (b) regenerating a plant from the transformed plant cell.

11. A method for producing a polynucleotide fragment comprising (a) selecting a nucleotide sequence comprised by the polynucleotide of claim 1, and (b) synthesizing a polynucleotide fragment containing the nucleotide sequence.

12. The method of claim 11, wherein the fragment is produced in vivo.

13. A chimeric gene comprising the polynucleotide of claim 1 operably linked to at least one regulatory sequence.

14. A method for altering the level of cysteine γ synthase expression in a host cell, the method comprising:(a) Transforming a host cell with the chimeric gene of claim 13; and(b) growing the transformed cell from step (a) under conditions suitable for the expression of the chimeric gene.

15. A method for evaluating a compound for its ability to inhibit the activity of a plant cysteine γ synthase, the method comprising the steps of:(a) transforming a host cell with a chimeric gene comprising a polynucleotide of claim 1, operably linked to at least one regulatory sequence;(b) growing the transformed host cell under conditions that are suitable for expression of the chimeric gene wherein expression of the chimeric gene results in production of the plant biosynthetic enzyme encoded by the operably linked nucleic acid fragment in the transformed host cell;(c) optionally purifying the plant biosynthetic enzyme polypeptide expressed by the transformed host cell;(d) treating the plant biosynthetic enzyme with a compound to be tested;(e) comparing the activity of the plant biosynthetic enzyme that has been treated with a test compound to the activity of an untreated plant biosynthetic enzyme polypeptide; and(f) selecting the compound that inhibits the activity of cysteine γ synthase.

16. An isolated polynucleotide comprising:(a) a nucleotide sequence encoding a polypeptide having plant amino acid biosynthetic activity, wherein the polypeptide has an amino acid sequence identity of at least 90% based on the Clustal method of alignment, when compared to a polypeptide selected from the group consisting of SEQ ID NOs: 2, 4, 6, 9, 11, 13, 15, 17, 19, 22, 24, 26, 28, 34, 36, 38, and 40, or(b) a complement of the nucleotide sequence, wherein the complement and the nucleotide sequence consist of the same number of nucleotides and are 100% complementary.

17. The isolated polynucleotide of claim 1, wherein the amino acid sequence of the polypeptide has at least 95% sequence identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs: 2, 4, 6, 9, 11, 13, 15, 17, 19, 22, 24, 26, 28, 34, 36, 38, and 40.

18. The isolated polynucleotide of claim 1, wherein the polynucleotide encodes a polypeptide selected from the group consisting of SEQ ID NOs: 2, 4, 6, 9, 11, 13, 15, 17, 19, 22, 24, 26, 28, 34, 36, 38, and 40.

19. The isolated polynucleotide of claim 1, wherein the polynucleotide comprises a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 8, 10, 12, 14, 16, 18, 21, 23, 25, 27, 33, 35, 37, and 39.

20. An isolated nucleic acid molecule that (1) comprises at least 180 nucleotides and (2) remains hybridized with a polynucleotide having a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 8, 10, 12, 14, 16, 18, 21, 23, 25, 27, 33, 35, 37, and 39 under a wash condition of 0.1.times.SSC, 0.1% SDS, and 65.degree. C.

21. A cell comprising the polynucleotide of claim 16.

22. The cell of claim 21, wherein the cell is selected from the group consisting of a yeast cell, a bacterial cell, and a plant cell.

23. A transgenic plant comprising the polynucleotide of claim 16.

24. A method for transforming a cell comprising introducing into a cell the polynucleotide of claim 16.

25. A method for producing a transgenic plant comprising:(a) transforming a plant cell with the polynucleotide of claim 16, and(b) regenerating a plant from the transformed plant cell.

26. A method for producing a polynucleotide fragment comprising:(a) selecting a nucleotide sequence comprised by the polynucleotide of claim 16, and(b) synthesizing a polynucleotide fragment containing the nucleotide sequence.

27. The method of claim 26, wherein the fragment is produced in vivo.

28. A chimeric gene comprising the polynucleotide of claim 16 operably linked to at least one regulatory sequence.

29. A method for altering the level of expression of a plant amino acid biosynthetic enzyme in a host cell, the method comprising:(a) transforming a host cell with the chimeric gene of claim 28; and(b) growing the transformed cell from step (a) under conditions suitable for the expression of the chimeric gene.

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