Biofilm Photobioreactor System And Method Of Use

Abstract

Flat panel biofilm photobioreactor systems with a photosynthetic, autofermentative microorganism that forms a biofilm and methods for using the same to make metabolic intermediate compound(s) through photosynthesis and to convert metabolic intermediate compound(s) into chemical product(s) such as a biofuel or a feedstock through autofermentation.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] Not applicable.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT

[0002] Not applicable.

REFERENCE TO SEQUENCE LISTING, A TABLE, OR A COMPUTER PROGRAM LISTING COMPACT DISC APPENDIX

[0003] This application includes a Sequence Listing submitted electronically as a text file named "Biofilm_Photobioreactor_System.txt", created on Sep. 28, 2012, with a size of 44 KB. The sequence listing consists of 16 sequences and is incorporated by reference into the specification in its entirety.

BACKGROUND

[0004] The possibility of using algae for the production of fuel and chemicals has attracted the interest of researchers, government and business for many years. Efforts to commercialize the production of fuel from algae have brought to light problems that must be solved to make this approach practical. The present invention is a novel approach to avoid or mitigate certain problems.

[0005] Under conventional approaches to algal biofuels, algal biomass is accumulated in open ponds or photobioreactors and harvested for conversion to fuel. The composition of the biomass may be altered to some extent through manipulation of the organism genetics or of the environment in which the organism is cultured, but generally there is a trade-off between optimizing composition and maximizing accumulation of biomass.

[0006] Under an alternative approach, the genetics and environment of a photosynthetic organism are manipulated to force the flux of carbon through photosynthesis into a desired product instead of toward accumulation of biomass. In the present invention, photosynthetic organisms are cultured as a biofilm inside a photobioreactor and maintained in stationary phase, and the environmental conditions in the photobioreactor are manipulated to induce the organisms to make a biofuel product, such as ethanol.

[0007] Obligate photosynthetic organisms require CO₂ as a feedstock to make a product or to accumulate biomass. One problem is that, if a photosynthetic organism is cultured in a body of water in a photobioreactor or pond that is exposed to CO₂ contained in air, then passive diffusion of carbon from air into water across the gas/liquid interface generally is not as efficient in sustaining maximal rates of photosynthesis. Accordingly, supplemental CO₂ usually must be added to cultures of photosynthetic organisms that are highly productive. The use of supplemental CO₂ significantly increases capital costs and operating costs, thereby reducing the profitability and rate of return of CO₂-supplemented systems.

[0008] Limited diffusion may be ameliorated by increasing the exposure of the culture of photosynthetic organisms to air, beyond the degree of exposure found with a culture contained in a body of water. Diffusion of CO₂ into a culture increases as the surface area of the culture that is in contact with air increases. In addition, many organisms express carbonic anhydrase at the cell surface, which catalyzes the conversion of carbon dioxide and water to bicarbonate and protons and thereby increases the rate of diffusion of CO₂ from the gas phase into the liquid phase:

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[0009] As a result, an air-exposed culture of an appropriate organism can achieve high productivity without CO₂ supplementation.

[0010] In many circumstances, gas phase delivery of CO₂ is preferred over other delivery modes. Because the concentration of CO₂ in air is typically less than 0.1% by volume, sustained elevated productivity by a culture requires considerable air throughput. The present invention provides a photobioreactor with a short airflow path and sufficiently low resistance to airflow that the necessary throughput of air can be achieved without high capital or operating costs. In some circumstances, the most economical delivery of CO₂ may be from a concentrated source instead of from the air. In some circumstances the most economical delivery of CO₂ may be as a solution of bicarbonate. The present invention provides a photobioreactor compatible with all of these modes of CO₂ delivery.

[0011] Another problem is that highly productive photosynthetic cultures tend to accumulate oxygen, which is a product of oxygenic photosynthesis. High oxygen concentrations in the culture can reduce productivity both by competing for photosynthetically produced electrons and through the effects of oxygen toxicity.

[0012] In a conventional photobioreactor containing a liquid suspension culture, oxygen may be removed from the culture by vigorous gas sparging, but high energy costs may be involved. This consideration militates in favor of gas phase exchange a preferred method of removing excess oxygen from the culture. The present invention facilitates gas phase exchange by providing a very short diffusion path for oxygen removal from the culture to the air stream.

[0013] Another problem is that an organism that channels photosynthetic energy primarily into making a fuel or chemical product, instead of accumulating biomass, severely disadvantaged compared to a competing organism that does not make the fuel product, and instead channels photosynthetic energy toward growth. This disparity reduces the stability of a culture of organisms that make a fuel product, since the culture may be invaded and outcompeted by other species that do not make the fuel product. Also, the organism ill undergo mutations that reduce the tendency to make the fuel product, thereby conferring a selective advantage over the productive, non-mutant type. As a result, the non-productive mutants will take over the culture, reducing or eliminating the productive organisms. This problem can be mitigated if production of the fuel or chemical product is beneficial to the organism.

[0014] If the organism makes the fuel product through fermentation, then production of the fuel product is necessary for the metabolism of the organism under anaerobic conditions and consequently the culture is more stable against mutation of the organism or invasion by non-fermenting species. The present invention facilitates fermentation to make a fuel product.

[0015] Product stability can be problematic if the fuel or chemical product is present in the culture and oxygen is present in the culture. The growth of aerobic heterotrophic bacteria that consume the product and that are present in the culture as contaminants is enabled by the availability of both oxygen and the product.

[0016] To address this problem, conventional photobioreactor or pond cultures must either incorporate unbreachable sterility barriers or must use antibiotics or other means so they are tolerant of some degree of contamination. The present invention minimizes the effect of contamination by heterotrophs on product stability and net productivity by substantially removing the product so that it is not present when oxygen is present in the culture.

[0017] Toxic effects of products such as ethanol on the organism of interest may also present problems. While product toxicity increases with productivity and product concentration, product toxicity can be mitigated by limiting the duration of exposure of the organism to the maximum product concentration. Further, exposure of the culture to product can be limited to the fermentation period, which may be conducted in darkness, in order to avoid toxicity responses that result from an interaction with photosynthetic processes.

[0018] Product purification costs are usually sensitive to concentration of the product that is extracted from a culture in a photobioreactor. It is desirable for fermentation to occur in a small fluid volume that yields elevated product concentration. The present invention provides a photobioreactor in which the fermentation volume is very small.

[0019] Capital costs must be kept within reasonable bounds for a fuel production system or method to be economically feasible. Materials, construction methods and supporting infrastructure must be chosen or designed with low cost in mind. A system of the present invention can have low material costs and a simplified infrastructure, and may be made using simple construction methods suitable for mass production. A system of the present invention may be light weight, minimizing mounting costs.

[0020] Because photosynthetic organisms in photobioreactors require exposure to sunlight, the culture in a photobioreactor may be exposed to high temperatures that are inimical to culture health and productivity. The present invention allows the management of culture temperature at low cost.

[0021] The considerations outlined above illustrate that the productivity of organisms that are cultured in a photobioreactor to make biofuel through metabolic processes may be restricted severely by limitations on uptake of CO₂ by the culture, removal of oxygen from the culture, genetic stability of the culture, stability of the product made by the culture, toxicity effects of the product on the culture and temperature effects on the culture.

[0022] US 2009/0181434 A1 to Aikens et al. discloses transgenic bacteria engineered to accumulate carbohydrates and a photobioreactor for cultivating photosynthetic microorganisms comprising a non-gelatinous, solid cultivation support suitable for providing nutrients and moisture to photosynthetic microorganisms and a physical barrier covering at least a portion of the surface of the cultivation support.

[0023] Aikens does not provide for the possibility of anoxic fermentation in the reactor structure or mode of operation. The photobioreactor proposed by Aikens is very different in detail from the present invention, using a different medium delivery system, a different product harvest system, and a completely different mode of operation. It does have in common with the present invention the use of a photosynthetic biofilm. The advantages of the present invention are that (1) periodic immersion provides a much more reliable uniform hydration than water seeping or dripping from a header; (2) the complexity and cost of a reactor design of the present invention are much lower; and (3) a reactor design of the present invention design lends itself to easily establishing conditions suitable for fermentation.

[0024] US 2008/0160591 A1 to Wilson at al. discloses a photobioreactor system for production of photosynthetic microorganisms that includes the use of extended surface area and an external water basin. Wilson et al. is related to the present invention in that Wilson at al. teaches the use of plastic film and similar construction techniques to produce a pattern of heat sealed welds between opposite panels. This reflects the concern of Wilson at al. with reactor cost, which is a concern also addressed by the present invention. Wilson et al. provides a photobioreactor design that is suited for the cultivation of organisms suspended in water medium Wilson et al. is not suitable for cultivation of a photosynthetic biofilm, and hence it does not provide the separation of retained biomass from a secreted or soluble product, and it is not suitable for operation with an autofermentation cycle.

[0025] US 20090258404 A1 to Mikkelsen at al. discloses production of fermentation products such as ethanol and lactic acid in biofilm reactors by microorganisms immobilized on sterilized granular sludge. Mikkelsen et al. is similar to the present invention in that Mikkelsen at al. uses a biofilm and anoxic fermentation. The apparatus and method of Mikkelsen at al. are not suitable for a photosynthetic biofilm or for an alternation of photosynthesis and autofermentation conditions essential to the present invention.

[0026] U.S. Pat. No. 5,595,893 to Pometto at al. discloses a solid support made of a synthetic polymer for immobilization of microorganism cells to form a biofilm reactor or use in fermentation, in streams for bioremediation of contaminants, and in waste treatment systems. It is possible that the support specified by Pometto et al. would be useful in a photobioreactor of the present invention. The reactor design used by Pometto et al. and the method of use are not compatible with a photosynthetic biofilm and an alternation of photosynthesis and autofermentation conditions essential to the present invention.

[0027] These references do not teach an optimized biofilm photobioreactor system of he present invention that resolves the limitations discussed above.

SUMMARY

[0028] An object of the present invention is a photobioreactor system that supports environmental conditions in which suitable organisms form biofilms on support substrates inside the photobioreactor, make metabolic intermediate compounds through photosynthesis and convert the metabolic intermediates into chemical products such as biofuels or feedstocks through autofermentation. According to the present invention, the design of the photobioreactor system enables increased uptake of carbon dioxide by the biofilm, increased removal of oxygen improved genetic stability of the biofilm and improved stability of the chemical product, while mitigating toxicity effects of the chemical product on the biofilm and temperature effects on the biofilm. Further according to the present invention, the photobioreactor system advantageously maintains a low cost basis.

[0029] A biofilm photobioreactor system of the present invention comprises flexible film that defines a photobioreactor enclosure. The photobioreactor enclosure has a flat panel shape and at least one portion of the flexible film is translucent.

[0030] Channels defined by at least one partition are disposed in the photobioreactor enclosure. The channels are in fluid communication.

[0031] A suitable photosynthetic, autofermentative microorganism cultured to form a biofilm in the photobioreactor enclosure. The suitable microorganism makes one or more metabolic intermediate compounds during a photosynthesis phase and converts the metabolic intermediate compound(s) to chemical product(s) during an autofermentation phase

[0032] The suitable microorganism forms a biofilm on a support substrate. The support substrate is disposed in the channels and is fixed to the flexible film, the at least one partition forming the channels or a combination of both.

[0033] The photobioreactor enclosure of the biofilm photobioreactor system incorporates at least one port formed in the flexible film for adding and removing liquid and adding gas to the photobioreactor enclosure, and at least one gas exhaust vent formed in the flexible film.

[0034] A gas comprising carbon dioxide flows through the photobioreactor enclosure intermittently and contacts the biofilm during the photosynthesis phase. The gas enters the photobioreactor enclosure through the at least one port and leaves the photobioreactor enclosure through the gas exhaust vent.

[0035] A first liquid at least partially fills the photobioreactor enclosure intermittently and contacts the biofilm during the photosynthesis phase. The first liquid enters the photobioreactor enclosure and leaves the photobioreactor enclosure through the port.

[0036] A second liquid at least partially fills the photobioreactor enclosure and contacts the biofilm during the autofermentation phase. The second liquid enters the photobioreactor enclosure and leaves the photobioreactor enclosure through the port. The chemical product made by the biofilm during the autofermentation phase enters the second liquid

[0037] A further object of the present invention is a method of making a biofuel through providing carbon dioxide and light to a biofilm of a suitable organism that is cultured in a photobioreactor of the present invention, such that the biofilm makes a metabolic intermediate compound through photosynthesis, and then removing light and electron acceptors such as oxygen, such that the biofilm converts the metabolic intermediate compound into a chemical product through autofermentation.

[0038] In a method of the present invention, a biofilm comprising a suitable photosynthetic, autofermentative microorganism is exposed to light. The biofilm is disposed on a support substrate that is disposed in channels of a biofilm photobioreactor and is fixed to inner surfaces of the biofilm photobioreactor

[0039] The channels of the biofilm photobioreactor are at least partially filled alternately with a flow of gas comprising carbon dioxide and with a first liquid. The gas and the first liquid contact the biofilm and the suitable microorganism snakes a metabolic intermediate compound from light and carbon dioxide by photosynthesis.

[0040] The biofilm is deprived of light and the channels of the biofilm photobioreactor are at least partially filled with a second liquid, which contacts the biofilm and expels the gas from the biofilm photobioreactor through outlets. The suitable microorganism converts the metabolic intermediate compound(s) into chemical product(s) by autofermentation, and the chemical product(s) enters the second liquid. The second liquid containing the chemical product(s) may be extracted from the biofilm photobioreactor.

[0041] The foregoing and other features and advantages of the invention will become further apparent from the following detailed description of the presently preferred embodiments, read in conjunction with the accompanying drawings. The detailed description and drawings are merely illustrative of the invention, rather than limiting the scope of the invention being defined by the appended claims and equivalents thereof.

BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWING

[0042] Embodiments of the invention will be described below with reference to the following figures.

[0043] FIG. 1 shows a plan view of a flat panel biofilm photobioreactor design.

[0044] FIG. 2 shows a side sectional view of a flat panel biofilm photobioreactor design.

[0045] FIG. 3 shows a schematic diagram of a flat panel biofilm photobioreactor with gas and media supply.

[0046] FIG. 4 shows a block diagram of steps of operating a flat panel biofilm photobioreactor.

[0047] FIG. 5 shows a frequency diagram of steps of operating a flat panel biofilm photobioreactor.

[0048] FIG. 6 shows a plan view of a flat panel biofilm photobioreactor design with a partition.

[0049] FIG. 7 shows a plan view of a flat panel biofilm photobioreactor design with multiple channels for flow of air and water.

[0050] FIG. 8 shows a plan view of a flat panel biofilm photobioreactor design with multiple channels for flow of air and water.

[0051] FIG. 9 shows a perspective view of a flat panel biofilm photobioreactor design with multiple channels for flow of air and water.

[0052] FIG. 10 shows a perspective view of an array of at panel photobioreactors and a mounting system.

[0053] FIG. 11 shows organic acid production rates.

[0054] FIG. 12 shows organic acid production rates.

[0055] FIG. 13 shows final ethanol concentration achieved in a flat panel biofilm photobioreactor.

DETAILED DESCRIPTION

[0056] The present invention relates to biofilm photobioreactor systems and a method for using a biofilm-forming microorganism in a photobioreactor of the present invention to make metabolic intermediate(s) through photosynthesis and convert the metabolic intermediates) into chemical product(s) through autofermentation. The primary chemical product preferably is a biofuel, such as ethanol, hydrogen, propanol or butanol, or a chemical feedstock such as acetate, lactate or formate.

[0057] As used herein, the term "suitable organism" means a microorganism that is able to attach to and form a biofilm on a surface and also to make metabolic intermediate compounds through photosynthesis and chemical products such as biofuels or feedstocks through fermentation. Non-limiting examples of suitable organisms within the meaning of the present invention are Geitlerinema Lyngbya, Chroococcidiopsis, Calothrix, Cyanothece, Oscillatoria, Gloeothece, Microcoleus, Microcystis, Nostoc, Anabaena and Spirulina species. One of ordinary ski II in the art will recognize that other suitable organisms are within the scope of the present invention.

[0058] For commercial production of chemical products, it is preferable to use a suitable organism that produces only a single product. While such organisms are known, generally a mix of products is produced. It is possible, using classical genetics or genetic engineering techniques, to eliminate pathways that lead to undesired chemical products, thereby leaving only a pathway that leads to the desired chemical product. If the suitable organism does not produce the desired chemical product, the required genes can be introduced using genetic engineering techniques. It is possible in principal to begin with an organism that does not perform autofermentation and introduce the necessary genes to confer the ability to autoferment. The development of enhanced organisms is not essential to the utility of this invention, but the development of such organisms will clearly greatly enhance the commercial value of this invention.

[0059] As used herein, the term "biofilm" means an aggregate of suitable organisms in which cells adhere to each other on or within a surface, frequently embedded within a self-produced matrix of exopolysaccharide or extracellular polymeric substance (EPS). Formation of a natural biofilm begins with free-floating microorganisms attaching to a surface through real, reversible adhesion via van der Waals forces, followed by anchoring themselves sing cell adhesion structures such as pili. Biofilms are often found on solid substrates submerged in or exposed to an aqueous solution. Artificial biofilms can also be made, using a flocculating agent such as sodium silicate, or an immobilizing agent such as alginate.

[0060] As used herein, the terms "exopolysaccharide" and "extracellular polymeric substance" mean a polymeric conglomeration generally composed of extracellular polysaccharides and proteins that hold together and protect a biofilm in matrix form.

[0061] As used herein, the term "oxic" means a concentration of dissolved oxygen in water greater than about 30% saturation.

[0062] As used herein, the term "hypoxia" means a concentration of dissolved oxygen in water in the range of from about 1% to about 30% saturation.

[0063] As used herein, the term "anoxia" means a concentration of dissolved oxygen in water less than about 1% saturation.

[0064] As used herein, the term "anaerobic" refers to cellular metabolism in which oxygen is not used as an electron acceptor.

[0065] As used herein, the term "fermentation" means the process of extracting energy from metabolic intermediate compounds that are organic compounds, such as carbohydrates or osmoprotectants, under hypoxic or anoxic conditions without the use of a terminal electron acceptor such as oxygen, sulfate or nitrate. Fermentative microorganisms typically hydrolyze complex organic polymers (e.g., glycogen) to monomers (e.g., glucose), which are further converted to lower molecular weight organic acids and alcohols. For example, fermentation may include the process of glycolysis, in which glucose is metabolically converted into pyruvate, followed by conversion of pyruvate into ethanol The concentration of oxygen or another electron acceptor below which a suitable organism will begin fermenting depends on the metabolic profile of the particular organism.

[0066] As used herein, the term "dark fermentation" means the fermentation of metabolic intermediate compounds such as organic substrates to hydrogen in the absence of light.

[0067] As used herein, the term "autofermentation" means that metabolic intermediate compounds that are made anabolically by the cells during photosynthesis and stored internally are catabolized under hypoxic or anoxic conditions to yield energy that may be used by the cell. Without using supplemental organic compounds, autofermenting cells produce carbon dioxide, chemical products such as biofuels and feedstocks, and energy used to regenerate adenosine triphosphate.

[0068] Photobioreactors and methods in accordance with the present invention are useful for producing a wide range of chemical products including acids, alcohols, ketones and hydrogen, and more specifically chemical products such as ethanol, butanol, propanol, methanol, propanediol, butanediol, lactate, proprionate, acetate, succinate, butyrate, formate and acetone.

[0069] As used herein, the term "metabolic intermediate compound" means an organic compound made by a microorganism through photosynthesis. Non-limiting examples of metabolic intermediate compounds are carbohydrates and osmoprotectants.

[0070] As used herein, the term "carbohydrate" means an organic compound that consists only of carbon, hydrogen, and oxygen. Non-limiting examples of carbohydrates are glycogen and glucose.

[0071] As used herein, the term "osmoprotectant" means a small molecule that acts as an osmolyte and helps organisms survive osmotic stress. Non-limiting examples of osmoprotectants are trehalose and glucosyl-glycerol.

[0072] As used herein, the term "osmolyte" means a compound that plays a rode in maintaining fluid balance and volume of a microorganism cell.

[0073] As used herein, the term "chemical product" means an organic compound made by a microorganism through fermentation or autofermentation. Non-limiting examples of chemical products are biofuels and feedstocks.

[0074] As used herein, the term "biofuel" means a type of fuel that derives energy from biological carbon fixation. Biofuels include fuels derived from biomass conversion or from cell metabolism, as well as solid biomass, liquid fuels and various biogases. Biologically produced alcohols such as ethanol may be produced by the action of microorganisms and enzymes through the fermentation of sugars or starches. Non-limiting examples of biofuels are ethanol, hydrogen, propanol and butanol.

[0075] As used herein, the term "feedstock" means a chemical compound that can be used as the starting material to make other products of interest, where such products are made using means other than the biofilm photobioreactor of the present invention. Non-limiting examples of feedstocks are acetate, lactate and formate.

[0076] As used herein, the term "biofilm photobioreactor" means a device or system used to support a biologically active environment for the cultivation of photosynthetic, autofermentative microorganisms. The biofilm photobioreactor may be constructed of flexible film that may be translucent. A biofilm photobioreactor of the present invention may be semi-closed against the exchange of gases and contaminants with the outside environment while permitting penetration of light through walls of the biofilm photobioreactor, or otherwise incorporating a light source, to provide photonic energy input for the photosynthetic culture of microorganisms contained in the biofilm photobioreactor. Cells of the photosynthetic microorganism are immobilized in layers on a support substrate inside the biofilm photobioreactor and the cell layers accumulate over time, forming a biofilm.

[0077] As used herein, the term "flexible film" means a thin continuous polymeric material or coating. Non-limiting examples of materials that can be used in flexible films suitable for use with the present invention are polyolefins, polyesters and vinyl copolymers thereof.

[0078] As used herein, the term "translucent" means allowing light to pass through, with or without scattering of photons.

[0079] As used herein, the term "support substrate" means a surface upon which a suitable organism is able to adhere and form a biofilm, Non-limiting examples of substrates within the meaning of the present invention that may be used with suitable organisms to form biofilms are films, filters, fabrics, foams and felts of polyesters, polyolefins, polyurethanes, polyamides, polyimides, polycarbonates, polydienes and polyacrylics adhered to the plastic film of the photobioreactor. One of ordinary skill in the art will recognize that the use of other substrates is contemplated within the scope of the present invention.

[0080] As used herein, the term "sparging" means a process whereby a chemically inert gas is bubbled through a liquid.

[0081] As used herein, the term "electron acceptor" means a chemical entity that accepts electrons transferred to it from another compound. An electron acceptor is an oxidizing agent that by virtue of its accepting electrons, is itself reduced in the process. An electron acceptor can, be a chemical entity such as ferric iron, sulphate, nitrate or nitrite, for example. A terminal electron acceptor is a compound such as oxygen, that receives or accepts an electron during cellular respiration.

[0082] As used herein, the term "substantially depleted of electron acceptors" describes an environment in which the partial pressures of oxygen, nitrate, sulphate, ferric iron, nitrite and other electron acceptors are love enough to facilitate anaerobic metabolism.

[0083] As used herein, the term "medium" means a liquid or gel designed to support the growth of microorganisms.

[0084] As used herein, the term "BG-11" means a standard cultivation medium for cyanobacteria that is well known to those of skill in the art. BG-11 contains all of the nutrients required for growth of many species of cyanobacteria. BG-11 is sold by, for example, Sigma-Aldrich Co LLC as the product "Cyanobacteria BG-11 Freshwater Solution" under SKU C3061.

[0085] The term "about" is used herein to mean approximately, in the region of, roughly, or around. When the term "about" is used in conjunction with a numerical value/range, it modifies that value/range by extending the boundaries above and below the numerical value(s) set forth. In general, the term "about" is used herein to modify a numerical value(s) above and below the stated value(s) by a variance of 20%

[0086] FIGS. 1 and 2 show an embodiment of a biofilm photobioreactor 100 configured as a flat panel photobioreactor enclosure 110 with an internal void volume. The photobioreactor enclosure 110 is oriented more or less vertically, with a port 120 for inlet of gas flow 230 and a port 122 for inlet and outlet of liquid flow 240, and an exhaust vent 130 for gas flow 230. Gas flow 230 and liquid flow 240 through the inlet and outlet ports 120 and 122 may be controlled by valves, pumps 330, fans 190 or other suitable devices. The exhaust vent 130 for gas flow 230 may be passive, such that gas flow 230 inside the photobioreactor enclosure 110 is forced through the exhaust vent 130 by the pressure head inside the photobioreactor enclosure 110.

[0087] Positioning the inlet port 120 for gas flow 230 and the exhaust vents 130 near the top edge 180 of the photobioreactor enclosure 110 eliminates the need to use valves or other potentially costly devices that would be required to prevent liquid flow 240 out of the photobioreactor enclosure 110 through ports 120 positioned lower on the photobioreactor enclosure 110. Alternatively, the inlet port 120 for gas flow 230 can be positioned near the bottom edge 182 of the photobioreactor enclosure 110, with external protection against backflow. If desired, the photobioreactor enclosure 110 can have a single port 124 for both gas flow 230 and liquid flow 240, as shown in FIG. 8.

[0088] The biofilm photobioreactor 100 may be inoculated with a suitable organism that forms a biofilm 140 on a support substrate 150 inside the photobioreactor enclosure 110. The support substrate 150 may be fixed in place by adhering the support substrate 150 to at least one wall 160 or edge 180 of the photobioreactor enclosure 110. In some embodiments, one or more pieces of support substrate 154 extend vertically and horizontally between the edges 180 of the internal void volume of the photobioreactor enclosure 110. As depicted in FIGS. 1 and 2, the support substrate 150 lies underneath, and is covered by, the biofilm 140 growth.

[0089] In preferred embodiments, the photobioreactor enclosure 110 is made from flexible film. Exemplary properties of a suitable flexible film for use in the present invention are translucency, tolerance to UV radiation, low oxygen diffusivity, low cost, light weight and acceptable durability. Flexible film edges 180 are bonded together, preferably through heat sealing, to form the photobioreactor enclosure 110.

[0090] At least one flexible film wall 160 of the photobioreactor enclosure 110 is translucent so that the biofilm 140 on the support substrate 150 contained within the photobioreactor enclosure 110 may be exposed to light from the sun or another source that provides photosynthetically active radiation having wavelengths from 400 to 700 nanometers. If only one flexible film wall 160 of the photobioreactor enclosure 110 is translucent, then the support substrate 150 preferably is adhered to the non-translucent wall 160. Suitable organisms in the biofilm 140 utilize the light to make metabolic intermediate compounds through photosynthesis.

[0091] Operation of a biofilm photobioreactor 100 of the present invention may be understood with further reference to FIGS. 3, 4 and 5. The biofilm photobioreactor 100 is inoculated 340 with a suitable organism that forms a biofilm 140. Internal environmental conditions are established that allow the biofilm 140 to first conduct photosynthesis, then autofermentation.

[0092] To initiate a photosynthesis period 450, the biofilm 140 is exposed 350 to light, and carbon is supplied to the suitable organisms by contacting 360 a gas, such as air, comprising carbon dioxide with the biofilm 140. Gas flow 230 to the photobioreactor enclosure 110 can be supplemented with carbon dioxide, but preferably will be air that is not supplemented. Suitable organisms in the biofilm 140 utilize the carbon dioxide to make a metabolic intermediate compound through photosynthesis. The metabolic intermediate compound is stored in the biofilm 140.

[0093] Gas flow 230 through the photobioreactor enclosure 110 can be created by using, for example, a fan 190. Gas flow 230 enters the photobioreactor enclosure 110 through a port 120 and leaves the photobioreactor enclosure 110 through the exhaust vents 130. In some embodiments, gas flow 230 from a gas source 200 is moved to a gas manifold 210, which splits the gas flow 230 among multiple biofilm photobioreactors 100 configured in an array 220. One of skill in the art will appreciate that gas and liquid flows among the components shown in FIG. 3 can be established using tubing, hoses or other means comprising any materials suitable for accommodating gas and liquid flows.

[0094] The biofilm 140 will tend to become dehydrated as a result of exposure to gas flow 230 that contains less than 100% relative humidity, and as a result of exposure to light, which raises the temperature in the photobioreactor enclosure 110. Dehydration of the biofilm 140 will result in reduced productivity, so water, culture medium or another suitable liquid 250 is added 360 periodically to the photobioreactor enclosure 110 to immerse and hydrate the biofilm 140. In some embodiments, the liquid 250 is a freshwater medium. The photobioreactor enclosure 110 is filled with the liquid 250 to a level below the exhaust vents 130, which expels gas flow 230 from the photobioreactor enclosure 110.

[0095] During the photosynthesis period 450, gas flow 230 through the photobioreactor enclosure 110 is periodically alternated with liquid flow 240 that fills the photobioreactor enclosure 110 at least partially. In some embodiments, the liquid 250 is a photosynthesis medium 280 that is split by a manifold 270 among multiple biofilm photobioreactors 100 configured in an array 220.

[0096] The optimal time periods of gas flow 230 and liquid flow 240 can be determined experimentally by monitoring carbon dioxide concentration in the gas flow 230 exiting the photobioreactor enclosure 110 through the exhaust vents 130, which indicates the rate of consumption of carbon dioxide and the rate of productivity by the biofilm 140. Generally, liquid flow 240 to immerse the biofilm 140 should occur frequently enough to maintain a relatively constant rate of carbon dioxide consumption by the biofilm 140.

[0097] In some embodiments, each period of gas flow 230 continues for any length of time in the range of from about five minutes to about four hours. In some embodiments, the photobioreactor enclosure 110 is quickly filled with liquid 250 and then drained, with the total period of liquid flow 240 to fill and drain the photobioreactor enclosure 110 taking about 10 seconds. In some embodiments, the total period of liquid flow 240 continues for about 20 minutes, or for any period in the range of from about 10 seconds to about 20 minutes.

[0098] The photosynthesis period 450 may continue for the length of diurnal sunlight, or otherwise for any length of time that is sufficient for the biofilm 140 to make and accumulate metabolic intermediate compound(s). Subsequently, an autofermentation period 460 is initiated in order to force the biofilm 140 to convert the metabolic intermediate compound(s) into chemical product(s) through autofermentation.

[0099] The autofermentation period 460 may continue for the length of diurnal darkness, or otherwise for any length of time that is sufficient for the biofilm 140 to convert the accumulated metabolic intermediate compound(s) into chemical product(s). A highly active biofilm 140 may convert accumulated metabolic intermediate compound(s) in a shorter period of time. In some embodiments, the autofermentation period 460 continues for any period of time in the range of from about one hour to about 18 hours.

[0100] To induce an autofermentation period 460, the biofilm 140 is deprived 370 of light, and the photobioreactor enclosure 110 is at least partially filled 370 with liquid 250 to exclude gas flow 230 from the photobioreactor enclosure 110 and deprive the biofilm 140 of terminal electron acceptors such as oxygen. The suitable organisms that comprise the biofilm 140 produce oxygen when exposed to light, and the initiation of n autofermentation period 460 is hindered when the biofilm 140 is producing oxygen. Accordingly, depriving 370 the biofilm 140 of exposure to light facilitates the initiation of an autofermentation period 460.

[0101] In some embodiments, the liquid 250 is a fermentation medium 300 that is split by a manifold 290 among multiple biofilm photobioreactors 100 configured in an array 220.

[0102] The liquid 250 remains in the photobioreactor enclosure 110 for the duration of the autofermentation period 460. Gas flow 230 through the photobioreactor enclosure 110 during the photosynthesis period 450 is expelled through the exhaust vents 130 when the photobioreactor enclosure 110 is filled with the liquid 250 at the beginning of the autofermentation period.

[0103] FIG. 5 further exemplifies the sequence and relative duration of events in the operation of the biofilm photo bioreactor 100. Time increases from left to right along the horizontal scale. Light periods 390 and dark periods 400 of the diurnal cycle 402 are color-coded. The occurrence of "Photosynthesis Liquid Fill" 410, "Photosynthesis Liquid Drain" 420, "Gas Flow" 230, "Autofermentation Liquid Fill" 430 and "Autofermentation Liquid Drain" 440 are indicated by the placement of vertical lines or bars, with the height of each line or bar indicating the velocity of gas flow 230 or liquid flow 240 and the width of each line or bar indicating the duration of non-zero flow velocity. The opposition of "Photosynthesis Liquid Fill" 410 with "Photosynthesis Liquid Drain" 420, and "Autofermentation Liquid Fill" 430 with "Autofermentation Liquid Drain" 440 additionally indicate directionality of non-zero flow velocity. FIG. 5 accordingly shows that photosynthesis 450 occurs in the biofilm 140 during light periods 390, during which there are periods of gas flow 230 through the photobioreactor enclosure 110 alternating with shorter periods of liquid flow 240 filling and then draining the photobioreactor enclosure 110. The photobioreactor enclosure 110 is filled with liquid 250 for the duration of the dark periods 400, during which autofermentation 460 occurs.

[0104] After an autofermentation period 460 ends, the liquid 250 containing chemical product(s) may be removed 380 from the photobioreactor enclosure 110 and processed using suitable means known to one of skill in the art to extract chemical product(s) from the liquid. In some embodiments, liquid 250 containing chemical product(s) from multiple biofilm photobioreactors 100 configured in an array 220 is combined by a product manifold 310 and stored in a product reservoir 320. it is desirable to remove and store the liquid containing the chemical product(s) under anaerobic conditions in order to suppress the growth of heterotrophic microorganisms that may consume chemical product(s) and reduce yield.

[0105] The liquid 250 used to fill the photobioreactor enclosure 110 during the autofermentation period 460 may be a freshwater medium that is substantially depleted of terminal electron acceptors before it is added to the photobioreactor enclosure 110. In some embodiments, the medium used during an autofermentation period 460 contains a dissolved oxygen concentration of less than about 15% saturation.

[0106] When an autofermentation medium 300 is prepared, the concentration of dissolved oxygen may be reduced by sparging the autofermentation medium 300 with a gas such as nitrogen or by vacuum degasing the autofermentation medium 300, for example. The autofermentation medium 300 is preferably stored outside the photobioreactor enclosure 110 under anoxic conditions in order to prevent the absorption of oxygen.

[0107] As an alternative to using sparging or other pre-treatments, the concentration of dissolved oxygen and other terminal electron acceptors in the autofermentation medium 300 may be reduced by adding the autofermentation medium 300 to the photobioreactor enclosure 110 and using the biofilm 140 to consume residual or ambient oxygen and other terminal electron acceptors through respiration. The time required for the respiratory activity of the biofilm 140 to create hypoxic or anoxic conditions that are suitable for autofermentation will be determined by factors such as the respiratory rate of the biofilm 140 the volume of autofermentation medium 300 and the initial concentrations of oxygen and other electron acceptors in the autofermentation medium 300.

[0108] One of skill in the art will appreciate that the biofilm photobioreactor 100 can be appropriately designed by sizing the photobioreactor enclosure 110 and selecting the biofilm 140 organism and autofermentation medium 300 such that the biofilm 140 will consume residual or ambient oxygen and other terminal electron acceptors and begin an autofermentation period 460 within a practical time period. In some embodiments, creation of autofermentation conditions and conversion of accumulated metabolic intermediate compound(s) to the chemical product(s) may be accomplished by the biofilm photobioreactor 100 in about one hour.

[0109] It may also be desirable to minimize the concentration of terminal electron acceptors present in the liquid 250 that is used to moisten the biofilm 140 during the photosynthesis period 450. This treatment may help increase the efficiency and speed of the transition from photosynthesis period 450 to autofermentation period 460 by reducing the quantity of terminal electron acceptors that carries over from the photosynthesis period 450 to the autofermentation period 460. The optimal concentration of terminal acceptors in the liquid 250 during the photosynthesis period 450 would provide sufficient nutrients to support the productivity of the biofilm 146.

[0110] Exposure of the biofilm 140 to oxygen that diffuses into the photobioreactor enclosure 110 could retard the onset of the autofermentation period 460. To prevent oxygen diffusion, the photobioreactor enclosure 110 is constructed using flexible film that is substantially impermeable to oxygen. Film fabrication methods and film compositions that minimize oxygen diffusion are commercially available and known to those of ordinary skill in the art. Examples of substantially oxygen-impermeable flexible films that are commercially available are polyethylene, polyester, and barrier films such as 3M HB-P 69731, which comprises a polyester base film, a heat sealable ethylene vinyl acetate copolymer layer and a ceramic oxide coating.

[0111] The path of gas flow 230 through the photobioreactor enclosure 110 is primarily from the inlet part 120 to the exhaust vent 130, such that positioning the inlet port 120 and the exhaust vent 130 near the top edge 180 of the photobioreactor enclosure 110 may contribute to incomplete gas flow 230 through lower portions of the photobioreactor enclosure 110 and uneven distribution of carbon dioxide to the biofilm 140. Low concentrations of carbon dioxide in gas flow 230 used in the photobioreactor enclosure 110 may require delivery of large volumes of gas to the photobioreactor enclosure 110, resulting in significant energy costs. The combination of these effects may be mitigated by introducing one or more partitions 470 that channel gas flow 230 more evenly throughout the photobioreactor enclosure 110.

[0112] FIG. 6 shows an embodiment of a biofilm photobioreactor 100 that incorporates a partition 470 that directs gas flow 230 from the top edge 180 of the photobioreactor enclosure 110 toward the bottom edge 182, and back toward the top edge 180, where gas flaw 230 is expelled through the exhaust vent 130. The partition 470 and channels 480 may be created by the pattern of bonding plastic film in the body of the photobioreactor enclosure 110. This embodiment increases the uniformity of distribution of gaseous carbon dioxide in contact with the biofilm 140.

[0113] FIG. 7 shows an embodiment of the biofilm photobioreactor 100 with multiple partitions 470 creating multiple U-shaped channels 480, a gas port 120 along the top edge 180 of the photobioreactor enclosure 110 and a liquid port 122 along the bottom edge 182 of the photobioreactor enclosure 110. The ports 120 and 122 direct gas flow 230 and liquid flow 240 to each channel 480. In this embodiment, the biofilm 140 is disposed on support substrates 150 positioned in the channels 480. The opposing walls 160 of the photobioreactor enclosure 110 are sealed together in a pattern forming the channels 480.

[0114] Biofilm 140 formation in the longer U-shaped channels 480 illustrated in FIG. 7 may be unevenly more dense closer to the gas port 120 and less dense closer to the exhaust vents 130. This effect occurs because the suitable organisms in the biofilm 140 consume most of the carbon dioxide in the gas flow 230 exiting the gas port 120 and produce a denser biofilm 140 in that location, while concentration of carbon dioxide in the gas flow 230 close to the exhaust vents 130 is significantly decreased providing less feedstock for suitable organisms in that location. Additionally, entering gas flow 230 may have low water content, resulting in drying of the biofilm 140 close to the gas port 120.

[0115] FIG. 8 shows an embodiment of the biofilm photobioreactor 100 with multiple partitions 470 creating multiple linear channels 480 that are greater in number and have shorter path lengths for gas flow 230 compared with the embodiment shown in FIG. 7. A combined port 124 located along the bottom edge 182 of the photobioreactor enclosure 110 supplies gas flow 230 and liquid flow 240 to the channels 480.

[0116] A biofilm 140 is disposed on support substrates 150 positioned in the channels 480. The support substrate 150 may be adhered to a wall 160 and/or edge 180, 182, 184 or 186 of the photobioreactor enclosure 110. Alternatively, the support substrate 150 may be attached to and suspended between the partitions 470 that form the channels 480 in the photobioreactor enclosure 110.

[0117] Biofilm 140 formation in the embodiment shown in FIG. 8 is more uniform, since the distribution of gas and carbon dioxide to more channels 480 and the shorter path lengths for gas flow 230 minimize discrepancies in the concentration of carbon dioxide between the combined port 124 and the exhaust vents 130, allowing the microorganisms to consume carbon dioxide and form a biofilm 140 that is distributed more evenly along the height of each channel 480. The increased number of channels 480, shorter gas flow 230 path lengths and increased exposure of the biofilm 140 to gas flow 230 help to increase uptake of carbon dioxide by the biofilm 140 and increase the removal of oxygen from the biofilm 140. Gas flow 230 enters the channels 480 at the bottom, so that liquid draining from the upper portions of the biofilm 140 helps to keep the biofilm 140 hydrated near the combined port 124.

[0118] Pressure inside the photobioreactor enclosure 110 increases when gas flow 230 is added to the photobioreactor enclosure 110 and when the photobioreactor enclosure 110 is filled with liquid 250. The internal gas or liquid pressure causes the walls 160 to deform outward. There may be distortion of the photobioreactor enclosure 110 shape so that exterior areas of the photobioreactor enclosure 110 are no longer substantially flat but instead exhibit creases formed in the flexible film. Such creases introduce resistance in the path of gas flow 230 through the channels 480 and necessitate the use of higher pressure to maintain satisfactory gas flow 230. This effect is more pronounced with plastic film that is thicker and less flexible. Such distortion and creasing may be lessened or prevented by heat sealing the opposing walls 160 at intermittent points to prevent outward deformation under pressure.

[0119] The increase in total weight when the photobioreactor enclosure 110 is filled with liquid 250 may require the use of sturdier and potentially more expensive materials to construct the photobioreactor enclosure 110 and/or use of a mounting system 510. The biofilm photobioreactor 100 may need to be mounted on a frame. The strength required of the frame depends on the weight of the biofilm photobioreactor 100 and the amount of tension that develops when the photobioreactor enclosure 110 is filled with gas or liquid. The determination of suitable designs and identification of materials based on biofilm photobioreactor 100 weight, wind loading and tension is well known to those skilled in the art.

[0120] FIG. 9 illustrates an embodiment in which the photobioreactor enclosure 110 is attached to side supports 500. Exemplary side supports 500 are made of wood or any other material that is suitably rigid, lightweight and inexpensive. The photobioreactor enclosure 110 may be attached to side supports 500 by stapling any other suitable means of bonding or adhering.

[0121] Generally, a biofilm photobioreactor 100 of this design will retain its shape if it is held at the left and right edges 184 and 186 while the top and bottom edges 180 and 182 are left free. The internal pressure holds the channels 480 open and gives the structure sufficient stiffness to be self-supporting.

[0122] Photosynthetic microorganisms tend to operate best in a certain range of light intensity. Photosynthesis is most productive when the microorganisms are exposed to as much light as they can readily tolerate. Excessive light exposure can cause photoinhibition, leading to loss of productivity. Light exposure also raises the temperature in a photobioreactor enclosure 110, which can cause loss of efficiency or cell death.

[0123] The rate of gas flow 230 within the photobioreactor enclosure 119 can also be adjusted to provide cooling, for example by controlling the voltage of the power source for the fan 190. In order to avoid expenses associated with providing liquid and moving gas, it is preferable to use a microorganism that tolerates and performs well at elevated temperatures, but it is possible to use the aforementioned methods to reduce peak temperatures so the microorganism used need not withstand such extremes of temperature as would be the case without heat management.

[0124] A biofilm photobioreactor 100 must be mounted in a position that allows exposure of the biofilm 140 to light. In a preferred embodiment, a biofilm photobioreactor 100 set up outdoors is mounted so that its position can be adjusted on at least one axis to track the position of the sun and control exposure to incoming solar radiation and radiant energy input. For example, a biofilm photobioreactor 100 set up in the Northern Hemisphere can be mounted so that the translucent wall 160 of the photobioreactor enclosure 110 faces south and the angle of the biofilm photobioreactor 100 can be adjusted. Alternatively, the biofilm photobioreactor 100 can face east, and its angle can be adjusted as the sun moves during the day so that a more or less constant light intensity is maintained by controlling the angle with respect to the sun.

[0125] FIG. 10 shows a mounting system 510 for an array 220 of biofilm photobioreactors 100 that incorporates a motor 520 to control the positions of adjustment arms 530. The mounting system 510 and adjustment arms 530 permit the angle of the biofilm photobioreactors 100 to be adjusted in order to optimize light exposure and photobioreactor 100 temperature. The angular disposition of the array 220 of biofilm photobioreactors 100 can be controlled in parallel, so individual mechanical mounting and controls are not required.

[0126] The mounting system 510 may allow adjustment of the angle of the biofilm photobioreactor 100 such that the photobioreactor enclosure 110 can be made substantially horizontal when it is filled with liquid, in order to limit deformation of the shape of the photobioreactor enclosure 110. Alternatively or in addition, the biofilm photobioreactor 100 can be mounted on a frame that exerts tension on the biofilm photobioreactor 100 if the shape of the photobioreactor enclosure 110 deforms excessively.

[0127] The mounting system 510 allows vertical orientation of the biofilm photobioreactor 100, which increases culture surface area that is exposed to sunlight per reactor ground footprint area and the culture per volume is exposed to sunlight. Vertical orientation of the biofilm photobioreactor 100 enhances distribution of culture within the light field and may be used to optimize light adaption and utilization by the biofilm 140 through ensuring that the biofilm 140 is consistently exposed to the same amount of light at each position in the photobioreactor enclosure 110.

[0128] The degree of deformation of the photobioreactor enclosure 110 shape relates to internal pressure in the photobioreactor enclosure 110 and the width of the channels 480. In general, narrow channels 480 prevent the biofilm photobioreactor 100 walls 160 from deforming excessively. Very narrow channels 480 require a greater internal pressure to open.

[0129] The degree of deformation of the photobioreactor enclosure 110 also relates to the distance between the left and right edges 184 and 186 of the photobioreactor enclosure 110. The distance between the left and right edges 184 and 186 of the photobioreactor enclosure 110 is maximal when it is completely flat, so if the photobioreactor enclosure 110 is mounted with this maximal distance between its left and right edges 184 and 186, deformation will be largely prevented. If the photobioreactor enclosure 110 does not deform, the channel 480 cross-section will be very small and gas will not flow freely. Furthermore, if there is any irregularity in the photobioreactor enclosure 110, some channels 480 or parts of channels 480 may not open at all, resulting in failure of gas flow 230 and loss of productivity in the affected channels 480. As a result, there is an optimal spacing of the left and right edges 184 and 186 of the photobioreactor enclosure 110 so that channels 480 open reliably and allow uniform gas flow 230.

[0130] During cycles in which the photobioreactor enclosure 110 fills with liquid, channels 480 must also open enough to allow uniform flow. It is not desirable for the channels 480 to open excessively, which would necessitate pumping more liquid to fill the channels 480. Further, during the autofermentation period, it is desirable to have a small volume of liquid in the channels 480 so that the change in concentration of the chemical product in the liquid is relatively large.

[0131] Given these considerations, a biofilm photobioreactor 100 can be mounted so that the distance between the left and right edges 184 and 186 can be adjusted by exerting a pulling force that is perpendicular to the longitudinal axes of the chambers. Stretching the biofilm photobioreactor 100 laterally in this manner will decrease the depth and volume of the channels 480 such that liquid volume in the channels 480 is reduced and product concentration is increased.

[0132] This embodiment provides a variation in the method of operating the biofilm photobioreactor 100. At the beginning of the autofermentation period 460, the photobioreactor enclosure 110 may be stretched laterally to increase the distance between the left and right edges 184 and 186, after which the channels 480 of the biofilm photobioreactor 100 are at least partially filled with liquid 250. The reduction in volume of liquid 250 needed to fill the stretched channels 480 during autofermentation 460 increases concentration of the chemical product in the liquid 250, which can facilitate recovery of the chemical product from the liquid 250. The reduction in volume of liquid 250 also reduces the quantity of dissolved oxygen and other electron acceptors that must be consumed by the biofilm 140 before conditions for autofermentation exist, which can decrease the time needed for autofermentation to begin. The reduction in volume of liquid 250 also reduces the total weight of liquid 250 in the photobioreactor enclosure 110. Decreased liquid 250 weight permits the use of lighter materials for construction of the biofilm photobioreactor 100, which helps reduce capital costs, and also helps reduce operating costs for liquid 250 and energy consumption.

[0133] The volume of the photobioreactor enclosure 110 that is not occupied by the biofilm 140 and the support substrate 150 can be adjusted. When the photobioreactor enclosure 110 is filled with gas or liquid 250, the fluid exerts pressure on the walls 160 of the photobioreactor enclosure 110, forcing them outward so each channel 480 has a rounded configuration. This outward deformation of the walls 160 of the channels 480 results in lateral contraction of the photobioreactor enclosure 110, bringing its left and right edges 184 and 186 closer together. In contrast, because the partitions 470 between channels 480 run from top to bottom, there is very little change in the distance between the top and bottom edges 180 and 182 of the photobioreactor enclosure 110 when it is filled with fluid 250.

[0134] The structure of the photobioreactor enclosure 110, regardless of how it is mounted, sets an upper limit on the degree to which the channel 480 volume can be increased and the photobioreactor enclosure 110 width (distance between left and right edges 184 and 186) can be decreased. In practice, it is preferable to mount the photobioreactor enclosure 110 in such a way that the left and right edges 184 and 186 of the photobioreactor enclosure 110 are constrained so that the decrease in photobioreactor enclosure 110 width, and hence the increase in photobioreactor enclosure 110 volume, is limited.

[0135] The preferred volume for the photobioreactor enclosure 110 is not necessarily the same for all phases of its operation. In particular, the opening of the channels 480 after draining the liquid 250 and initiating gas flow 230 is most reliable when the photobioreactor enclosure 110 volume is at least about two liters per square meter of photobioreactor enclosure 110 surface area.

[0136] During the period of autofermentation 460, the chemical product concentration that can be reached in a single night depends on the productivity of the biofilm 140 and the liquid 250 volume. The desired chemical product concentration depends on the economics of purification and the tolerance of the microorganism to the chemical product. Chemical product concentration must be high enough that the chemical product can be economically recovered, but low enough to avoid an unacceptable level of stress to the organism.

[0137] As an example, assume that ethanol can be economically purified from liquid 250 that is at least 0.5% ethanol by weight, and the microorganism used is tolerant of nightly exposure to ethanol of at most 1%. If the nightly conversion of carbohydrate to ethanol is 10 grams of carbohydrate per square meter, or about 5.1 grams of ethanol per square meter, then the total liquid 250 volume should be in the approximate range of 500 ml to 1 liter.

[0138] Table 1 shows chemical product concentration as a function of productivity and liquid 250 volume.

TABLE-US-00001 TABLE 1 Total liquid volume per square meter 0.25 L 0.5 L 1 L 2 L 3 L 4 L 5 L Ethanol 2 g 0.41% 0.20% 0.10% 0.05% 0.03% 0.03% 0.02% productivity 5 g 1.02% 0.51% 0.26% 0.13% 0.09% 0.06% 0.05% per 10 g 2.04% 1.02% 0.51% 0.26% 0.17% 0.13% 0.10% square 15 g 3.07% 1.53% 0.77% 0.38% 0.26% 0.19% 0.15% meter 20 g 4.09% 2.04% 1.02% 0.51% 0.34% 0.26% 0.20% per day 25 g 5.11% 2.56% 1.28% 0.64% 0.43% 0.32% 0.26% 30 g 6.13% 3.07% 1.53% 0.77% 0.51% 0.38% 0.31% 35 g 7.16% 3.58% 1.79% 0.89% 0.60% 0.45% 0.36% 40 g 8.18% 4.09% 2.04% 1.02% 0.68% 0.51% 0.41%

[0139] Desirable chemical product concentration values preferably are in the range of from about 0.50% to about 1.00%. The chemical product concentration values presented in Table 1 are exemplary, based on reasonable ranges for ethanol production. Chemical product concentration values will differ for other chemical products, other microorganisms, and other separation technologies; suitable calculations can be made by one of ordinary skill.

[0140] The total liquid 250 volume includes both the added medium and the liquid 250 bound in the biofilm 140 and the support substrate 150, so the added medium will be smaller than the total liquid 250 volume. If the volume of liquid 250 in the biofilm 140 is a large fraction of the total liquid 250 volume, it may be necessary to include a wash step after the autofermentation period 460 to recover an acceptable fraction of the chemical product.

[0141] The liquid 250 volume can also be reduced by withdrawing liquid 250, which allows the channels 480 to collapse so that the opposing walls 160 contact each other. This normally occurs every time the photobioreactor enclosure 110 is drained. The channels 480 open when gas flow 230 resumes.

[0142] The reliability of channel 480 collapse depends on the material chosen for construction of the photobioreactor enclosure 110. Flexible films are preferred to allow for reliable channel 480 collapse.

[0143] The liquid 250 volume that remains after channel 480 collapse generally is smaller than the desired volume described above, so this method is most appropriate if the desired volume is unusually small. Alternatively, the photobioreactor enclosure 110 can be oriented approximately horizontally. The channels 480 will collapse when gas flow 230 ceases and can be filled with liquid 250 in this position, but the fill volume is considerably less than the fill volume of a vertical photobioreactor enclosure 110.

[0144] If a microorganism has a high tolerance for the chemical product, or if productivity is low, it may not be convenient to reduce the liquid 250 volume sufficiently to achieve an economically desirable concentration of chemical product in a single night of autofermentation. If this is the case, it is possible to reuse the autofermentation medium 300 several times so the chemical product concentration is increased to the desired level.

[0145] If a microorganism has a low tolerance for the chemical product, or if productivity is very high, it may not be convenient to increase the liquid 250 volume sufficiently to prevent damage to the microorganism. If this is the case, the liquid 250 can be withdrawn after a limited time period to avoid excess chemical product concentration and new medium can be added to continue autofermentation.

[0146] A separate advantage of a low fermentation volume is that the period of time required for the biofilm 140 to reduce the oxygen concentration sufficiently to induce autofermentation is decreased. For example, if an organism uses carbohydrate at the rate of 1 gram per square meter hour, it will consume oxygen at a rate of about 1 gram per square meter hour. A 1 liter volume will only have about 10 milligrams of oxygen, so the time to anoxia in this example is less than 1 minute.

[0147] In most cases, the considerations noted previously are more significant than time to anoxia in determining the optimal water volume. Time to anoxia may be important if it is necessary to use large volumes and respiratory rates are low. Time to anoxia may also be reduced by pretreatment of the fermentation medium by degassing, for example by vacuum degassing or heat degassing, by chemical oxygen scavenging, by sparging, for example with nitrogen, or by biological processes such as bacterial growth in a closed container without aeration.

[0148] When autofermentation medium is stored, either between fermentations that reuse the same medium or if it is necessary to hold the product containing medium before it goes on to a product separation step, the fermentation medium can be held in conditions in which the amount of oxygen that can enter the stored medium is insufficient to allow a significant amount of biological degradation of the product to occur, for example in a closed tank with a limited or zero headspace.

[0149] Internal pressure stresses portions of flexible film that are sealed together, such as partitions 470, and potentially can induce mechanical failure. Accordingly, the film used to construct the photobioreactor enclosure 110, the size of the channels 480 and the operating pressure of the photobioreactor enclosure 110 must be selected to ensure that mechanical failure does not occur.

[0150] Another design consideration is that ports 120, 122 and 124 must be sized to avoid excessive head loss during periods of gas and liquid movement.

[0151] The biofilm photobioreactor 100 incorporates a control system to operate pumps 330, fans 190 and similar equipment for the purpose of adding and removing gas and liquid to and from the photobioreactor, as well as to operate motors 520 used to adjust a mounting system 510 if it is adjustable. The control system can use inputs such as wind speed, air temperature and light intensity to adjust the angle of the photobioreactors to achieve optimal biofilm photobioreactor 100 performance with respect to photosynthetic rate, biofilm photobioreactor 100 temperature and avoidance of mechanical failure due to wind loading.

[0152] It is advantageous for the biofilm 140 to have a high specific rate of fermentation. The rate of fermentation can be affected by medium constituents. Generally, the medium used during photosynthesis will be chosen to facilitate high photosynthetic efficiency and photosynthate accumulation, while the fermentation medium will be chosen to facilitate high fermentation rate. The fermentation medium will also be chosen to facilitate extraction of the chemical product from the medium.

[0153] The rate of fermentation is also determined by the characteristics of the microorganism in the biofilm 140 and its specific fermentation rate. Generally, fermentation rate in autotrophic microorganisms is related to the energy demands of the microorganism and the availability of food reserves. In order to obtain the greatest productivity for a given biomass, it is desirable for the specific fermentation rate to be high.

[0154] The process of fermentation can adversely affect the medium and cause accumulation of chemical products that are toxic to the biofilm 140 at elevated concentrations. Depending on the fermentation rate and the tolerance of the microorganism for accumulated chemical product(s), the liquid used during autofermentation may be extracted periodically from the photobioreactor enclosure 110 to harvest chemical product(s), and then the photobioreactor enclosure 110 may be refilled with fresh liquid. For example, if a biofilm 140 measuring 1 square meter has accumulated 20 grams of carbohydrate that it ferments to ethanol in 500 milliliters of water, then the resulting ethanol concentration will be slightly over 2%. If the organism tolerant of exposure to 1% ethanol but not 2%, then it is necessary to extract the medium containing the ethanol.

[0155] Medium in which chemical product is collected may be used in more than one autofermentation period before it is processed to extract the chemical product. The medium stored under anaerobic conditions prior to processing in order to protect the accumulated chemical product from aerobic heterotrophic microorganisms. After the medium is processed and the chemical product is extracted, the medium may be reused in successive autofermentation periods.

[0156] Autofermentation will not necessarily occur during every dark period.

[0157] Instead of adding autofermentation liquid, the dark cycle can comprise flowing gas at a low flow rate through the photobioreactor enclosure 110, alternated with periodic, brief submersion of the biofilm 140 to assure sufficient hydration, similar to the daytime cycle. For example, if accumulation of metabolic intermediate compound(s) in the biofilm 140 is only 5 grams per square meter per day, the autofermentation medium volume is 500 milliliters of water, and the microorganism can tolerate 2% ethanol, then it is desirable to permit up to 4 days of accumulation of metabolic intermediate compound(s) before autofermentation is induced so that the autofermentation medium will reach the tolerance limit of the microorganism. Achieving a higher ethanol concentration before harvesting the ethanol is desirable because higher product concentration reduces purification costs.

[0158] Autofermentation also does not need to occur during the initial growth of the biofilm 140. Gas flow 230 may be maintained during dark conditions at a low rate with periodic immersion if necessary for hydration. Once the biofilm 140 is mature and ready to be productive, chemical product can be harvested by inducing autofermentation.

Example 1

[0159] Two biofilm photobioreactors, PBR 1 and PBR 2, were fabricated out of 3M Scotchpak HB-P 69731 Translucent High Barrier Film. This film was chosen due to its resistance to oxygen permeability and ease of use in prototype construction.

[0160] The photobioreactor enclosures each incorporated a gas port positioned near the top edge of the photobioreactor enclosure, a liquid port positioned near the bottom edge of the photobioreactor enclosure and partitions creating four "U" shaped channels, similar to the design shown in FIG. 7. Ambient air was introduced into the channels via the gas port using an aquarium pump with flow capacity of 2.25-4.50 liters per minute. The channels were fashioned so that air entered into the "U" via the gas port, flowed down through the "U" and then back up to an 18-20 gauge gas exhaust vent. Media were introduced and removed via the liquid port.

[0161] Support substrate fabric of woven polyester was seamed into each photobioreactor enclosure and acted as a substrate for biofilm development. Total surface area of each photobioreactor enclosure was 0.1428 square meters and the volume of each photobioreactor enclosure was 0.415 liters.

[0162] Operation of the biofilm photobioreactors consisted of a photosynthesis cycle during which glycogen was produced and stored, and an autofermentation cycle during which glycogen was catabolized. In photosynthesis cycles, a freshwater photosynthesis medium was pumped through the liquid port, filling the photobioreactor enclosure, and then the medium was immediately pumped through the liquid port from the photobioreactor enclosure to a storage reservoir.

[0163] In the autofermentation cycle, a nitrogen-sparged, freshwater autofermentation medium was pumped through the liquid port, filling the photobioreactor enclosure, and remained in the photobioreactor enclosure until just before the beginning of the photosynthesis cycle. The autofermentation medium was depleted of SO₄ and other terminal electron acceptors. No air was introduced into the photobioreactor enclosures during the autofermentation period. The process of regularly exchanging media and delivering air to the photobioreactors was automated using proprietary software that controlled peristaltic pumps and air pumps.

[0164] PBR 1 and PBR 2 were each inoculated with 15 mL culture of Chroococcidiopsis sp. into a 500 ml reservoir of marine BG-11, made with filtered seawater containing about 1-3% dissolved salts. The biofilm photobioreactors were mounted in front of cool white fluorescent lamps providing about 75 μmol photons per square meter per second at each photobioreactor surface. The photobioreactor enclosures were filled and immediately drained every 20 minutes.

[0165] Regular photosynthesis cycles were initiated every 30 minutes. The biofilm photobioreactors were maintained at about 27° C. with a 24-hour photoperiod at an irradiance of 75 μmol photons per square meter per second. The BG-11 medium was incrementally changed to the freshwater photosynthesis medium over the course of 12 days. After the media exchange was completed, light was adjusted to a 16-hr photoperiod.

[0166] Autofermentation cycles were initiated. The autofermentation cycles and nitrogen sparging were automated to provide nitrogen gas flow to the autofermentation medium reservoir for 1.5 hours before the autofermentation cycle was initiated by flooding the photobioreactor enclosure with the autofermentation medium. After several hours, the autofermentation medium was pumped out of the photobioreactor enclosure, and the photosynthesis cycle resumed. Automated autofermentation cycles occurred each day, with the photobioreactor enclosure placed in darkness through shading with black cloth.

[0167] After the autofermentation medium was pumped into each photobioreactor enclosure, initial samples for organic acids, ethanol, oxygen and pH were taken by gravity draining a small autofermentation medium aliquot from the photobioreactor enclosure. Organic acid samples (in duplicate) were filtered through a 0.2 μm syringe filter and stored at -80° C. until analysis. Ethanol samples (in duplicate) were aliquoted into gas chromatography (GC) vials and stored at -20° C. until analysis. Organic acid and ethanol samples were quantified less than 7 days from sampling. Dissolved oxygen and pH measurements were taken immediately using bench top probes. At the end of the autofermentation cycle, final samples were obtained as described above. Prior to automation, photobioreactors were sampled each time an autofermentation cycle was attempted. Once the photobioreactors were automated, sampling occurred three times per week.

TABLE-US-00002 TABLE 2 Autofermentation Cycle Data for PBR 1 and PBR 2 Beginning of Autofermentation Cycle End of Autofermentation Cycle PBR Cycle Cycle O₂ Lactate Acetate Formate O₂ Lactate Acetate Formate No. (hrs) Number pH (mg/L) (ppm) (ppm) (ppm) pH (mg/L) (ppm) (ppm) (ppm) 1 6.5 1 0.86 1.0 7.2 0.2 0.58 12.0 24.8 1.8 1 6.2 2 0.23 <0.2 1.1 <0.2 0.13 2.0 4.3 0.2 1 21.5 3 0.71 <0.2 <0.2 <0.2 0.65 14.6 21.4 2.5 2 21.5 4 0.66 <0.2 <0.2 <0.2 0.49 20.5 24.7 2.6 1 4.5 5 0.42 <5 <5 -- 9.17 0.42 5.7 7.3 -- 2 4.5 6 0.39 <5 -- -- 9.13 1.44 <5 <5 -- 1 5.5 7 7.65 0.64 <5 <5 -- 6.92 0.76 7.2 9.8 -- 2 5.5 8 9.11 0.65 <5 <5 -- 7.90 0.63 <5 5.9 -- 1 6 9 7.32 0.50 <5 -- -- 6.50 1.05 8.7 7.3 -- 2 6 10 9.06 0.75 -- -- -- 7.91 1.42 <5 5.8 --

[0168] Autofermentation cycle length varied from 4.5 to 21.5 hours. Oxygen ranged from 0.23 to 0.86 mg/L at the start of the autofermentation cycle. Initial concentrations of organic acids, lactate, acetate, and formate ions were very low or below detection limits in the medium at the beginning of the autofermentation cycle. At the end of the autofermentation cycles, oxygen ranged from 0.13 to 1.44 mg/liter. Organic acids were present at the end of the autofermentation cycle in all but one experimental trial, confirming autofermentation by Chroococcidiapsis sp. in biofilm photobioreactors. Organic acid yields, expressed as concentrations at the autofermentation cycle end, were highest in the longest duration cycle. Medium pH was variable but declined following autofermentation.

[0169] Acetate was typically the most abundant organic acid produced during autofermentation, although both lactate and acetate were present at the autofermentation cycle end with concentrations ranging from 2 to 20.5 ppm. Formate was only detected in three trials in very low concentrations.

[0170] Assuming that concentrations reported as below the quantification limits ere effectively zero, organic acid production rates as a function of illuminated culture surface area (single sided illumination) were calculated using the following equation:

P=(C_final-C_initial)/t×PBR_volume/PBR_illuminated area Eq. (1)

where P is the production rate, C_final and C_initial are the final and initial organic acid concentrations, PBR_volume is the biofilm photobioreactor volumetric capacity, PBR_illuminated area is the biofilm photobioreactor surface area that is exposed to light (single-sided illumination) and t is time. Organic acid production rates were similar, with average values of 3.26 mg per square meter per hour for lactate and 3.93 mg per square meter per hour for acetate (FIG. 11). Formate production was extremely low,

[0171] Ethanol was measured on each occasion and was not detected in any sample.

Example 2

[0172] Two biofilm photobioreactors, PBR 3 and PBR 4, were fabricated out of 3M Scotchpak HB-P 69731 Translucent High Barrier Film. Each photobioreactor enclosure incorporated a combination port for gas and liquid flow positioned near the bottom edge of the photobioreactor enclosure and partitions creating eight "l" shaped channels, similar to the design shown in FIG. 8, Ambient air was introduced into each channel via the combination port using an aquarium pump with flow capacity of 2.25-4.50 liters per minute. Air entered each channel from the bottom through the combination port and flowed up to an 18-20 gauge gas exhaust vent Media entered each channel and were removed via the combination port.

[0173] Support substrate fabric of woven polyester was seamed into each photobioreactor enclosure and acted as a substrate for biofilm development. Total surface area of each photobioreactor enclosure was 0.1428 square meters and the volume of each photobioreactor enclosure was 0.415 liters.

[0174] Operation of the biofilm photobioreactors consisted of a photosynthesis cycle during which glycogen was produced and stored, and an autofermentation cycle during which glycogen was catabolized. Photosynthesis cycles were initiated by flushing each photobioreactor enclosure with freshwater photosynthesis medium. After the photobioreactor enclosures were filled and immediately drained through the combination port using the freshwater photosynthesis medium, ambient air was pumped through each photobioreactor enclosure using aquarium air pumps. Air exchange was the primary method of oxygen management. The freshwater photosynthesis medium flush occurred every 30 minutes during the 18-hr photosynthesis cycle.

[0175] Autofermentation cycles were initiated by turning off air flow to the photobioreactor enclosures, then pumping nitrogen-sparged, freshwater autofermentation medium into the photobioreactor enclosures through the combination port. The autofermentation medium was depleted of SO₄ and other terminal electron acceptors. The autofermentation medium was held in each photobioreactor enclosure for the duration of the 6-hr autofermentation cycle. The process of regularly exchanging media and delivering air to the photobioreactor enclosures was automated using proprietary software that controlled peristaltic pumps and air pumps.

[0176] PBR 3 and PBR 4 were each inoculated with 500 mL culture of Geitlerinema sp. into a 500 ml reservoir of marine BG-11. The biofilm photobioreactors were mounted in front of cool white fluorescent lamps providing about 75 μmol photons per square meter per second at each photobioreactor surface. The photobioreactor enclosures were filled and immediately drained every 30 minutes.

[0177] The biofilm photobioreactors were maintained at about 27° C. with a 16-hour photoperiod at an irradiance of 75 μmol photons per square meter per second. The marine BG-11 medium was incrementally changed to the freshwater photosynthesis medium over the course of 16 days.

[0178] Daily six-hour autofermentation cycles were initiated. One 24-hour autofermentation cycle was also completed,

[0179] After the autofermentation medium was pumped into each photobioreactor enclosure, initial samples for organic acids, ethanol, oxygen and pH were taken by gravity draining a small autofermentation medium aliquot from the photobioreactor enclosure. Organic acid samples (in duplicate) were filtered through a 0.2 μm syringe filter and stored at -80° C. until analysis. Ethanol samples (in duplicate) were aliquoted into gas chromatography (GC) vials and stored at -20° C. until analysis. Organic acid and ethanol samples were quantified less than 7 days from sampling. Dissolved oxygen and pH measurements were taken immediately using bench top probes. At the end of the autofermentation cycle, final samples ere obtained as described above.

[0180] Four autofermentation cycles were attempted in each of PBR 3 and PBR 4 (Table 3). During a 24-hour autofermentation cycle in PBR 4, the photobioreactor system developed a leak in the pump tubing and autofermentation medium drained from the photobioreactor enclosure. Consequently, no data are presented in Table 3 for that cycle.

TABLE-US-00003 TABLE 3 Autofermentation Cycle Data for PBR 3 and PBR 4 Beginning of Autofermentation Cycle End of Autofermentation Cycle PBR Cycle Cycle O₂ Lactate Acetate Formate O₂ Lactate Acetate Formate No. (hrs) Number pH (mg/L) (ppm) (ppm) (ppm) pH (mg/L) (ppm) (ppm) (ppm) 3 6 1 9.82 2.71 1.05 1.01 -- 9.06 1.0 0.78 12.51 0.44 4 6 2 9.15 1.05 1.22 0.95 -- 8.58 1.28 7.27 33.21 1.90 3 6 3 9.74 0.96 1.06 1.04 -- 9.13 0.81 3.28 10.20 0.88 4 6 4 9.43 0.83 0.78 1.02 -- 8.82 1.20 3.67 28.73 0.88 3 24 5 9.53 1.21 -- 1.32 -- 8.55 0.4 2.14 21.77 4.62 3 6 6 9.11 1.24 0.98 -- 1.26 8.66 0.81 -- -- 3.05 4 6 7 9.02 0.96 3.20 -- 1.36 8.51 0.78 1.29 2.51 3.52

[0181] Production rates (standardized to illuminated area) were highest for acetate, with the exception of cycle 6 (FIG. 12). Lactate consumption was observed in cycles 1, 6 and 7. Formate production rates averaged 0.52 mg per square meter per hour ±0.27 mg per square meter per hour and 0.80 mg per square meter per hour ±0.33 mg per square meter per hour, respectively, for PBR 3 and PBR 4. Production rates for organic acids were higher in PBR 4 for all autofermentation cycles.

[0182] Ethanol was not detected in any samples at the start of any of the autofermentation cycles. However, low concentrations of ethanol were measured at the end of autofermentation cycles 1, 2, 4, 5 and 7 (FIG. 13). Final concentrations of ethanol in PBR 3 and PBR 4 ranged from 0.0002 to 0.0020% v/v.

Example 3

[0183] SEQ ID NO: 1-16 disclosed herein identify DNA sequences and protein sequences for a Geitlerinema sp. that is similar to the Geitlerinema sp. described in Examples 1 and 2, wherein the DNA sequences and protein sequences encode for enzymes that potentially are used to make ethanol from pyruvate through autofermentation.

TABLE-US-00004 TABLE 4 SEQ ID NO Sequence Type Enzyme 1 DNA Pyruvate dehydrogenase, E1-α 2 Encoded protein Pyruvate dehydrogenase, E1-α 3 DNA Pyruvate dehydrogenase, E1-β 4 Encoded protein Pyruvate dehydrogenase, E1-β 5 DNA Pyruvate dehydrogenase, E2 6 Encoded protein Pyruvate dehydrogenase, E2 7 DNA Alcohol dehydrogenase 8 Encoded protein Alcohol dehydrogenase 9 DNA Bifunctional alcohol dehydrogenase 10 Encoded protein Bifunctional alcohol dehydrogenase 11 DNA Acetyl-CoA synthetase 12 Encoded protein Acetyl-CoA synthetase 13 DNA Acetaldehyde dehydrogenase 14 Encoded protein Acetaldehyde dehydrogenase 15 DNA Acetatekinase 16 Encoded protein Acetatekinase

[0184] While the embodiments of the invention disclosed herein are presently considered to be preferred, various changes and modifications can be made without departing from the scope of the invention. The scope of the invention is indicated in the appended claims, and all changes that come within the meaning and range of equivalents are intended to be embraced therein.

Sequence CWU 1

1

1611128DNAGeitlerinema sp. 1ttgattgccg gtcgatcggt ctcatcagac acattggggc aactcgtccc gtatcctata 60tttattaagt taaagtttgc aacgtcgaaa ctaatggtca cagaacgcac aatccccacg 120tcctacagca atacggctga aatcagccgg gaagaagggc tgcgagtcta cgaagacatg 180gttttagggc gcttcttcga ggacaagtgc gccgaaatgt actatcgcgg taaaatgttc 240ggcttcgttc acctctacaa cggtcaagaa gccgtctcct ccggggtcat ccaagccatg 300cgccccggcg acgattacgt ttgcagcacc taccgcgacc acgttcacgc tctcagttgc 360ggtgttcccg cccgtgaagt gatggcggaa ctgttcggaa aatcgaccgg atgcagtaaa 420gggcgcgggg ggtcgatgca tatgttctcg gaaccccacc gactcctcgg cggttatgcg 480tttgtcgctg aaggtattcc cgtggcgatg ggggcggcgt tccaagtgcg ctatcgcaag 540gaagtcatgg gcgattcctc cgccgaccaa gtggtggcct gcttcttcgg cgacggtgcc 600agtaacaacg gtcaattttt cgagaccttg aacatggcgt cgttgtggaa gctaccgatt 660attttcgtcg tcgaaaacaa taaatgggcg atcgggatgg ctcacgaccg ggcgacctcg 720caaccggaga tttacaaaaa agccagcgtg tttaacatgg ctggggtcga agtggatgga 780atggacgtga tggcggtgcg ggcggcagcc caagaggcgg tcgatcgcgc gcgggccgga 840gaaggcccca cgctcatcga ggcgttgacg taccgcttcc ggggacactc tctcgccgac 900ccggacgaac tgcgatcgaa agaggagaag gaaatctggt tctctcgcga cccgattcac 960cgctttgaaa actacctgac ggaagaaaac ctcgccagtg cggaggaact caaggacatt 1020cagaagaaaa ttcaagaggt catcgacgac tcggtggaat tcgccgaatc cagccccgaa 1080cccgacccca gcgaacttcg tcgctttatc tttgcagaag acgaataa 11282375PRTGeitlerinema sp. 2Leu Ile Ala Gly Arg Ser Val Ser Ser Asp Thr Leu Gly Gln Leu Val 1 5 10 15 Pro Tyr Pro Ile Phe Ile Lys Leu Lys Phe Ala Thr Ser Lys Leu Met 20 25 30 Val Thr Glu Arg Thr Ile Pro Thr Ser Tyr Ser Asn Thr Ala Glu Ile 35 40 45 Ser Arg Glu Glu Gly Leu Arg Val Tyr Glu Asp Met Val Leu Gly Arg 50 55 60 Phe Phe Glu Asp Lys Cys Ala Glu Met Tyr Tyr Arg Gly Lys Met Phe 65 70 75 80 Gly Phe Val His Leu Tyr Asn Gly Gln Glu Ala Val Ser Ser Gly Val 85 90 95 Ile Gln Ala Met Arg Pro Gly Asp Asp Tyr Val Cys Ser Thr Tyr Arg 100 105 110 Asp His Val His Ala Leu Ser Cys Gly Val Pro Ala Arg Glu Val Met 115 120 125 Ala Glu Leu Phe Gly Lys Ser Thr Gly Cys Ser Lys Gly Arg Gly Gly 130 135 140 Ser Met His Met Phe Ser Glu Pro His Arg Leu Leu Gly Gly Tyr Ala 145 150 155 160 Phe Val Ala Glu Gly Ile Pro Val Ala Met Gly Ala Ala Phe Gln Val 165 170 175 Arg Tyr Arg Lys Glu Val Met Gly Asp Ser Ser Ala Asp Gln Val Val 180 185 190 Ala Cys Phe Phe Gly Asp Gly Ala Ser Asn Asn Gly Gln Phe Phe Glu 195 200 205 Thr Leu Asn Met Ala Ser Leu Trp Lys Leu Pro Ile Ile Phe Val Val 210 215 220 Glu Asn Asn Lys Trp Ala Ile Gly Met Ala His Asp Arg Ala Thr Ser 225 230 235 240 Gln Pro Glu Ile Tyr Lys Lys Ala Ser Val Phe Asn Met Ala Gly Val 245 250 255 Glu Val Asp Gly Met Asp Val Met Ala Val Arg Ala Ala Ala Gln Glu 260 265 270 Ala Val Asp Arg Ala Arg Ala Gly Glu Gly Pro Thr Leu Ile Glu Ala 275 280 285 Leu Thr Tyr Arg Phe Arg Gly His Ser Leu Ala Asp Pro Asp Glu Leu 290 295 300 Arg Ser Lys Glu Glu Lys Glu Ile Trp Phe Ser Arg Asp Pro Ile His 305 310 315 320 Arg Phe Glu Asn Tyr Leu Thr Glu Glu Asn Leu Ala Ser Ala Glu Glu 325 330 335 Leu Lys Asp Ile Gln Lys Lys Ile Gln Glu Val Ile Asp Asp Ser Val 340 345 350 Glu Phe Ala Glu Ser Ser Pro Glu Pro Asp Pro Ser Glu Leu Arg Arg 355 360 365 Phe Ile Phe Ala Glu Asp Glu 370 375 3984DNAGeitlerinema sp. 3atggcagaaa ccttattttt caacgcttta agagaagcca tcgacgaaga aatggcgcga 60gacgagaccg tcttcgtcct cggagaggat gtcggtcact acggcggttc ctacaaagtt 120accaaagacc tctatcaaaa atacggcgaa ctgcgcctgc tcgacacgcc catcgccgaa 180aacagtttca cggggatggc cgtaggtgcg gcgattacag ggttgcgacc catcatcgaa 240ggcatgaaca tggggtttct gctcctggcc ttcaaccaaa tcgccaataa tgccgggatg 300ttgcgctaca cctccggcgg aaacttcaaa attcccatgg tgattcgcgg tccgggtggc 360gtcggacgcc aactcggtgc cgaacactct caacggctcg aagcgtactt tcaagccgtt 420ccggggttga aaatcgtcgc ttgttcgacc ccgtacaacg cgaaaggttt gctcaaagcc 480gctattcgcg acaacaaccc agtattgttc ttcgaacacg tgctgctcta caacctcaaa 540gaaaacttac ccgagagcga atacgtcgtt cccctcgata aagccgaagt ggtgcgagac 600ggaaaagacg tgacgatttt gacctactcg cggatgcgcc accactgcac gcaagcggca 660aaaaccttag aaaaagacgg gttcgatccc gaaattatcg acttgatttc cctcaagccc 720tacgacctcg aaaccatcgg aaattcgatt cgcaaaaccc accgcgttat cgtcgtcgaa 780gaatgtatga aaactggcgg tgtcggtgcc gaactgatcg ccaccatcaa cgaccatttc 840ttcgacgaac tcgacgcccc cgtgattcgt ttgtcctccc aagacattcc gacgccctat 900aacggaatgc tcgaacggtt gacgatcgtg caaccgcatc aaatcgtcga agcggttcag 960aacatggtgg cgttaaaagt gtag 9844327PRTGeitlerinema sp. 4Met Ala Glu Thr Leu Phe Phe Asn Ala Leu Arg Glu Ala Ile Asp Glu 1 5 10 15 Glu Met Ala Arg Asp Glu Thr Val Phe Val Leu Gly Glu Asp Val Gly 20 25 30 His Tyr Gly Gly Ser Tyr Lys Val Thr Lys Asp Leu Tyr Gln Lys Tyr 35 40 45 Gly Glu Leu Arg Leu Leu Asp Thr Pro Ile Ala Glu Asn Ser Phe Thr 50 55 60 Gly Met Ala Val Gly Ala Ala Ile Thr Gly Leu Arg Pro Ile Ile Glu 65 70 75 80 Gly Met Asn Met Gly Phe Leu Leu Leu Ala Phe Asn Gln Ile Ala Asn 85 90 95 Asn Ala Gly Met Leu Arg Tyr Thr Ser Gly Gly Asn Phe Lys Ile Pro 100 105 110 Met Val Ile Arg Gly Pro Gly Gly Val Gly Arg Gln Leu Gly Ala Glu 115 120 125 His Ser Gln Arg Leu Glu Ala Tyr Phe Gln Ala Val Pro Gly Leu Lys 130 135 140 Ile Val Ala Cys Ser Thr Pro Tyr Asn Ala Lys Gly Leu Leu Lys Ala 145 150 155 160 Ala Ile Arg Asp Asn Asn Pro Val Leu Phe Phe Glu His Val Leu Leu 165 170 175 Tyr Asn Leu Lys Glu Asn Leu Pro Glu Ser Glu Tyr Val Val Pro Leu 180 185 190 Asp Lys Ala Glu Val Val Arg Asp Gly Lys Asp Val Thr Ile Leu Thr 195 200 205 Tyr Ser Arg Met Arg His His Cys Thr Gln Ala Ala Lys Thr Leu Glu 210 215 220 Lys Asp Gly Phe Asp Pro Glu Ile Ile Asp Leu Ile Ser Leu Lys Pro 225 230 235 240 Tyr Asp Leu Glu Thr Ile Gly Asn Ser Ile Arg Lys Thr His Arg Val 245 250 255 Ile Val Val Glu Glu Cys Met Lys Thr Gly Gly Val Gly Ala Glu Leu 260 265 270 Ile Ala Thr Ile Asn Asp His Phe Phe Asp Glu Leu Asp Ala Pro Val 275 280 285 Ile Arg Leu Ser Ser Gln Asp Ile Pro Thr Pro Tyr Asn Gly Met Leu 290 295 300 Glu Arg Leu Thr Ile Val Gln Pro His Gln Ile Val Glu Ala Val Gln 305 310 315 320 Asn Met Val Ala Leu Lys Val 325 51323DNAGeitlerinema sp. 5atgatccacg aaatcttcat gcctgcccta agttccacca tgactgaagg caaaatcgtc 60tcttggacga aatccccagg ggacaaggtg gaaaaaggcg aaacggtggt tgtcgtcgag 120tcggacaaag ctgacatgga cgtggagtcc ttctacgaag ggattctggc gaccatcgtt 180gtcggagaag gcgacgtcgc ccccgtcggc ggaaccatcg ccctgttagc cgaaacggaa 240gcggaaatcg aagaagctaa gcaaaaagcc cagcaacagc aacaagggca accccaaacg 300gcggccgctt cggaaacgcc gtcaaccccc caaccgactc cagccgcagc gacggcacaa 360aacggcgcgt ctcaagcggc ggccagcgga caaaatggcg gacgcattgt cgcctctcct 420cgcgcccgga agttagccaa agagctgaaa gtcgatttga acgggttgca cgggagcggt 480ccttacggtc gtatcgtggc tgaagacgtg caagcagcgg ccggacaacc cgtcctaccg 540actgcaacgg cagttgcacc gatgccctca gctcccgctc cgtcggcagt tccctcggtt 600ccggcgcaag cccccacgac aacggctccg gctgccacta gcgccccctt gggtcaggtg 660gttccgttca atacgctgca aggggcagtg gtgcggaaca tgacggcgag tttgcaagtg 720ccgacgttcc acgtgggtta caccatcacc acggacaatt tagacgcctt ataccagcaa 780attaagtcca aaggtgtgac gatgacgggg ctactggcga aagccgtggc ggtgacgttg 840cagaaacacc cgttacttta cgccagctac accgaacagg gcgttcagta caacagcaac 900attaacgtgg cggtggcggt ggccatgccc ggaggcggct tgattacgcc ggtgatgcgc 960gatgccgacc agatggatat ttactcgctg tctcggtcgt ggaaggatct cgtggcgcga 1020tcgcgtgcca aacaactgca accggaagag tacagcacgg gaacctttac cctgtccaat 1080ttgggaatgc tcggggtcga tcgcttcgat gcgattttac cccccggaca aggatcgatt 1140ctcgcgatcg gtgcctctcg tccccaggtt gtcgccacgg acgacggcat gatgggcgtc 1200aaacgtcaaa tgcaagtgaa cattacctgc gaccaccgca ttatttacgg tgcagacgcg 1260gcggcgttct tgaaagattt ggccgacctc gttgaaaaca acccgcaatc tctaacgctg 1320taa 13236440PRTGeitlerinema sp. 6Met Ile His Glu Ile Phe Met Pro Ala Leu Ser Ser Thr Met Thr Glu 1 5 10 15 Gly Lys Ile Val Ser Trp Thr Lys Ser Pro Gly Asp Lys Val Glu Lys 20 25 30 Gly Glu Thr Val Val Val Val Glu Ser Asp Lys Ala Asp Met Asp Val 35 40 45 Glu Ser Phe Tyr Glu Gly Ile Leu Ala Thr Ile Val Val Gly Glu Gly 50 55 60 Asp Val Ala Pro Val Gly Gly Thr Ile Ala Leu Leu Ala Glu Thr Glu 65 70 75 80 Ala Glu Ile Glu Glu Ala Lys Gln Lys Ala Gln Gln Gln Gln Gln Gly 85 90 95 Gln Pro Gln Thr Ala Ala Ala Ser Glu Thr Pro Ser Thr Pro Gln Pro 100 105 110 Thr Pro Ala Ala Ala Thr Ala Gln Asn Gly Ala Ser Gln Ala Ala Ala 115 120 125 Ser Gly Gln Asn Gly Gly Arg Ile Val Ala Ser Pro Arg Ala Arg Lys 130 135 140 Leu Ala Lys Glu Leu Lys Val Asp Leu Asn Gly Leu His Gly Ser Gly 145 150 155 160 Pro Tyr Gly Arg Ile Val Ala Glu Asp Val Gln Ala Ala Ala Gly Gln 165 170 175 Pro Val Leu Pro Thr Ala Thr Ala Val Ala Pro Met Pro Ser Ala Pro 180 185 190 Ala Pro Ser Ala Val Pro Ser Val Pro Ala Gln Ala Pro Thr Thr Thr 195 200 205 Ala Pro Ala Ala Thr Ser Ala Pro Leu Gly Gln Val Val Pro Phe Asn 210 215 220 Thr Leu Gln Gly Ala Val Val Arg Asn Met Thr Ala Ser Leu Gln Val 225 230 235 240 Pro Thr Phe His Val Gly Tyr Thr Ile Thr Thr Asp Asn Leu Asp Ala 245 250 255 Leu Tyr Gln Gln Ile Lys Ser Lys Gly Val Thr Met Thr Gly Leu Leu 260 265 270 Ala Lys Ala Val Ala Val Thr Leu Gln Lys His Pro Leu Leu Tyr Ala 275 280 285 Ser Tyr Thr Glu Gln Gly Val Gln Tyr Asn Ser Asn Ile Asn Val Ala 290 295 300 Val Ala Val Ala Met Pro Gly Gly Gly Leu Ile Thr Pro Val Met Arg 305 310 315 320 Asp Ala Asp Gln Met Asp Ile Tyr Ser Leu Ser Arg Ser Trp Lys Asp 325 330 335 Leu Val Ala Arg Ser Arg Ala Lys Gln Leu Gln Pro Glu Glu Tyr Ser 340 345 350 Thr Gly Thr Phe Thr Leu Ser Asn Leu Gly Met Leu Gly Val Asp Arg 355 360 365 Phe Asp Ala Ile Leu Pro Pro Gly Gln Gly Ser Ile Leu Ala Ile Gly 370 375 380 Ala Ser Arg Pro Gln Val Val Ala Thr Asp Asp Gly Met Met Gly Val 385 390 395 400 Lys Arg Gln Met Gln Val Asn Ile Thr Cys Asp His Arg Ile Ile Tyr 405 410 415 Gly Ala Asp Ala Ala Ala Phe Leu Lys Asp Leu Ala Asp Leu Val Glu 420 425 430 Asn Asn Pro Gln Ser Leu Thr Leu 435 440 7875DNAGeitlerinema sp. 7atggcaaaga ttcaggctta tgctgcccga gacgtggcgg gaaaactcga accgtttgaa 60tacgatcccg gtgcgttgga tgccgaagag gtcgaactcg ctgtcgaatc ctgcgggatc 120tgtcacagcg acctgagtat gttagacgac gaatgggaga tgacgcaata tcccttcgtt 180cccggtcacg aagtcgtcgg aaccgtgacg gagatcggcg atcgcgtcac cgatctccaa 240atcggacagc gggtcggttt gggctggttt gccaactcct gtatgggttg ccagtggtgt 300atgtccggcg accacaatct ctgtagcgac gccgaaggca cgatcgtcgg acgtcacggc 360ggatttgccg atcgcgttcg cgcccatcac agttgggtca tcccgattcc cgagggcatc 420gatcccctca aggccgggcc gctgttctgc ggtggcatta ccgtattcaa tccgatggta 480gagttcgacc tcaaacccac cgatcgcgtg ggtgtggtgg gaattggcgg cctcggacac 540ctggcgattc aattcctcag tgcttgggga tgcgaggtga cggcgttctc gacgagtgcg 600gacaaggaag ccgaagccaa agaactcggg gcgacccatt ttgtcaattc taaagatctc 660gacgccctgc aagcggtcga aaactccttt gattttatta tttcgacggt gagtgccgat 720ctcgattgga acgcttacgt ggcggcgttg cgaccgaagg gacgattgca cttagttggc 780gtggcgacga atcccctcga tttacagttg tttcctctgc tgatgggtca gaagtcggtg 840tcgtcgagtc cggtggggag tccggtgacg atcgc 8758291PRTGeitlerinema sp. 8Met Ala Lys Ile Gln Ala Tyr Ala Ala Arg Asp Val Ala Gly Lys Leu 1 5 10 15 Glu Pro Phe Glu Tyr Asp Pro Gly Ala Leu Asp Ala Glu Glu Val Glu 20 25 30 Leu Ala Val Glu Ser Cys Gly Ile Cys His Ser Asp Leu Ser Met Leu 35 40 45 Asp Asp Glu Trp Glu Met Thr Gln Tyr Pro Phe Val Pro Gly His Glu 50 55 60 Val Val Gly Thr Val Thr Glu Ile Gly Asp Arg Val Thr Asp Leu Gln 65 70 75 80 Ile Gly Gln Arg Val Gly Leu Gly Trp Phe Ala Asn Ser Cys Met Gly 85 90 95 Cys Gln Trp Cys Met Ser Gly Asp His Asn Leu Cys Ser Asp Ala Glu 100 105 110 Gly Thr Ile Val Gly Arg His Gly Gly Phe Ala Asp Arg Val Arg Ala 115 120 125 His His Ser Trp Val Ile Pro Ile Pro Glu Gly Ile Asp Pro Leu Lys 130 135 140 Ala Gly Pro Leu Phe Cys Gly Gly Ile Thr Val Phe Asn Pro Met Val 145 150 155 160 Glu Phe Asp Leu Lys Pro Thr Asp Arg Val Gly Val Val Gly Ile Gly 165 170 175 Gly Leu Gly His Leu Ala Ile Gln Phe Leu Ser Ala Trp Gly Cys Glu 180 185 190 Val Thr Ala Phe Ser Thr Ser Ala Asp Lys Glu Ala Glu Ala Lys Glu 195 200 205 Leu Gly Ala Thr His Phe Val Asn Ser Lys Asp Leu Asp Ala Leu Gln 210 215 220 Ala Val Glu Asn Ser Phe Asp Phe Ile Ile Ser Thr Val Ser Ala Asp 225 230 235 240 Leu Asp Trp Asn Ala Tyr Val Ala Ala Leu Arg Pro Lys Gly Arg Leu 245 250 255 His Leu Val Gly Val Ala Thr Asn Pro Leu Asp Leu Gln Leu Phe Pro 260 265 270 Leu Leu Met Gly Gln Lys Ser Val Ser Ser Ser Pro Val Gly Ser Pro 275 280 285 Val Thr Ile 290 91587DNAGeitlerinema sp. 9ttgatgggta ttgcagcaga gaaactcgcc ccaatttcga tcgtgttgtt tgccgccgga 60attttaatct gggggtatta ccgcgctcga ccctacggca aaatcggctt gctgtcttgg 120ttgcagtcgg tttctgtcac agctccctgg ctgctgtttt tcgccctgtt ctcagccggg 180attttcatca atttcgcagg cgtgttgttc ctcatcgtcg cgtccgtttt aacttacatc 240gggctgggac gacaactgcg agcggcggct cgagatcccg aacaacgagc gtacttagaa 300aaactcgcga acgcccaacg gtcttcaaaa tctgacgagt cctctcaaac ttccgatctt 360gccgaaccgt ccgaagactc tgaagcggcc gaaccgtctc cggacaccgc acgaccgacg 420gagccagttg ccgcctctgt ccgagatcgc gatcgagaaa gcgatcgccc gtttcagaac 480ctatccgttc cggaggacga cttacactgc attcaagaaa tcttcggaat cgatacgttt 540ttcgccacag agacgatccc gtaccagtcg ggagcaattt tcaaaggcaa ccttcgggga 600gaggtcgaag cgacccatca ggaactgtcg aaaaagctgc acgatcgcgt gggcgatcgc 660taccgtttgt ttttcgtcaa cgatcccgac gaaaaaacgg tcgtcgtcgt cttgccgagt 720cgtaacgacc ctcaaccgct cacgacgaac caacagattt tagcggtcgt attgttcgtc 780gcgacgatcg tcaccactct cgaaaccggg ggggcatttc tcggattcga cttgttcgag 840aatttaaacc gctggacgga aacactgccc ctcgccttgg gaatttgggc gatattgctc 900gttcacgaac tcggccaccg tatcgcggcc ggacgatatg gcattgcgct ctcgccgcca 960tttttccttc ccacctggca aatcggctct ttcggtgcga ttacccgttt cgagtcgttg 1020ctgcccaacc gttcgaccct gttcgacatc gccatcgccg

ggccggccgc cggtgggttg 1080ttgtccttgg gaatgttagc ggtcgggttc gttctgtccc acgacgggag tttgtttcaa 1140cttccgagcg aatttttccg agggtcggtt ttggtgggat tgctggccaa ggcgttttta 1200ggtgaagccc tccagcagag tttggtagac gtgcatccgt tggtcgttct cggttggctg 1260gggttagtca ttaacgccct caacctgatt ccagccggac agttagacgg cgggcgcgtc 1320atgcaggcga tttacggtcg tcggattgcg gggcgatcga cgatcgccac cctcatcgtg 1380ttagcgatcg cttccttcgt caatccgtta gccttgtact gggcgatcgt gattttggtc 1440atccaacggg atttagagcg tccgagtctc aacgaaatta ccgaacccga cgacacccgt 1500gcaattttgg cgttcgtggc actgttggtc atgttgatga cgttgattcc ctttacgccg 1560agtttggcgt tgcgcctggg gctgtaa 158710528PRTGeitlerinema sp. 10Leu Met Gly Ile Ala Ala Glu Lys Leu Ala Pro Ile Ser Ile Val Leu 1 5 10 15 Phe Ala Ala Gly Ile Leu Ile Trp Gly Tyr Tyr Arg Ala Arg Pro Tyr 20 25 30 Gly Lys Ile Gly Leu Leu Ser Trp Leu Gln Ser Val Ser Val Thr Ala 35 40 45 Pro Trp Leu Leu Phe Phe Ala Leu Phe Ser Ala Gly Ile Phe Ile Asn 50 55 60 Phe Ala Gly Val Leu Phe Leu Ile Val Ala Ser Val Leu Thr Tyr Ile 65 70 75 80 Gly Leu Gly Arg Gln Leu Arg Ala Ala Ala Arg Asp Pro Glu Gln Arg 85 90 95 Ala Tyr Leu Glu Lys Leu Ala Asn Ala Gln Arg Ser Ser Lys Ser Asp 100 105 110 Glu Ser Ser Gln Thr Ser Asp Leu Ala Glu Pro Ser Glu Asp Ser Glu 115 120 125 Ala Ala Glu Pro Ser Pro Asp Thr Ala Arg Pro Thr Glu Pro Val Ala 130 135 140 Ala Ser Val Arg Asp Arg Asp Arg Glu Ser Asp Arg Pro Phe Gln Asn 145 150 155 160 Leu Ser Val Pro Glu Asp Asp Leu His Cys Ile Gln Glu Ile Phe Gly 165 170 175 Ile Asp Thr Phe Phe Ala Thr Glu Thr Ile Pro Tyr Gln Ser Gly Ala 180 185 190 Ile Phe Lys Gly Asn Leu Arg Gly Glu Val Glu Ala Thr His Gln Glu 195 200 205 Leu Ser Lys Lys Leu His Asp Arg Val Gly Asp Arg Tyr Arg Leu Phe 210 215 220 Phe Val Asn Asp Pro Asp Glu Lys Thr Val Val Val Val Leu Pro Ser 225 230 235 240 Arg Asn Asp Pro Gln Pro Leu Thr Thr Asn Gln Gln Ile Leu Ala Val 245 250 255 Val Leu Phe Val Ala Thr Ile Val Thr Thr Leu Glu Thr Gly Gly Ala 260 265 270 Phe Leu Gly Phe Asp Leu Phe Glu Asn Leu Asn Arg Trp Thr Glu Thr 275 280 285 Leu Pro Leu Ala Leu Gly Ile Trp Ala Ile Leu Leu Val His Glu Leu 290 295 300 Gly His Arg Ile Ala Ala Gly Arg Tyr Gly Ile Ala Leu Ser Pro Pro 305 310 315 320 Phe Phe Leu Pro Thr Trp Gln Ile Gly Ser Phe Gly Ala Ile Thr Arg 325 330 335 Phe Glu Ser Leu Leu Pro Asn Arg Ser Thr Leu Phe Asp Ile Ala Ile 340 345 350 Ala Gly Pro Ala Ala Gly Gly Leu Leu Ser Leu Gly Met Leu Ala Val 355 360 365 Gly Phe Val Leu Ser His Asp Gly Ser Leu Phe Gln Leu Pro Ser Glu 370 375 380 Phe Phe Arg Gly Ser Val Leu Val Gly Leu Leu Ala Lys Ala Phe Leu 385 390 395 400 Gly Glu Ala Leu Gln Gln Ser Leu Val Asp Val His Pro Leu Val Val 405 410 415 Leu Gly Trp Leu Gly Leu Val Ile Asn Ala Leu Asn Leu Ile Pro Ala 420 425 430 Gly Gln Leu Asp Gly Gly Arg Val Met Gln Ala Ile Tyr Gly Arg Arg 435 440 445 Ile Ala Gly Arg Ser Thr Ile Ala Thr Leu Ile Val Leu Ala Ile Ala 450 455 460 Ser Phe Val Asn Pro Leu Ala Leu Tyr Trp Ala Ile Val Ile Leu Val 465 470 475 480 Ile Gln Arg Asp Leu Glu Arg Pro Ser Leu Asn Glu Ile Thr Glu Pro 485 490 495 Asp Asp Thr Arg Ala Ile Leu Ala Phe Val Ala Leu Leu Val Met Leu 500 505 510 Met Thr Leu Ile Pro Phe Thr Pro Ser Leu Ala Leu Arg Leu Gly Leu 515 520 525 111849DNAGeitlerinema sp. 11atgtcagaac agacgataga atcaattctt cacgagcaac ggacgtttcc cccagccgca 60gattttgccg ctaacgccca tatcaaaagc atggccgact ataaggcttt gtgcgatcgc 120gccgaaaaag acccggctgg attttggagc gaactggccg aaaccgaact cgactggttt 180caaaagtggg agaacgtcct cgactggcaa ccccccgttg ccaaatggtt cgagggaggc 240aaactcaacg tttcttacaa ctgcctcgat cgccatctga ccacctggcg caaaaacaaa 300gccgcgttga tttgggaagg ggaacctggc gactcgcgca ccctcaccta cgcccaactg 360caccgcgaag tgtgccagat ggccaacgtc atcaaacagt tcggcgtgaa aaaaggcgat 420gtcgtgggga tttatatgcc catgattccc gaagcggcga tcgccatgtt agcctgcgcc 480cgcatcggtg ccgttcacag cgtcgtgttc ggtggcttca gtgccgaagc cctgcgcgat 540cgcgtcaacg ccgccgaagc caaactggtg attaccgccg acggcggctt ccgcaaagac 600aaagtcgtca ccctcaaaga ccaagtcgat aaagccctcg ccaacgacgc cgcccccagc 660gtcgaaaacg ttctcgtggt gcgtcgcatc gaaaaagaca ctcacatgga agagggtcgg 720gaccactggt ggcacgaagt ccgtcaaggc atctccgccc actgtcctgc cgaaccgatg 780gacagcgaag acatcctttt catcctctac accagcggca gcaccggaaa accgaaaggc 840gtcgttcaca ccaccgccgg atacaacctc tacgcccacg tcaccaacaa atggacgttc 900gacctgaagg acaccgatat tttctggtgt accgccgacg tgggttggat taccggacac 960agctatatcg tctacggtcc gctgtctaac ggggcgacga cggtgatgta cgaaggggtt 1020ccccgtccgt ccaaccccgg ctgtttttgg gatgtcgtgg aaaaatacgg cgtcacgatt 1080ttctacaccg cccccaccgc cattcgtgcc tttattaaag ccggagacaa acacccgaac 1140gcccgcgatt tgtccagctt gcggctgttg ggaaccgtgg gcgaacccat caacccgaaa 1200gcctggatgt ggtatcaccg agtcatcggc ggcgaacgct gtccggtcgt cgatacctgg 1260tggcagacgg aaacgggcgg tttcatgatt acgccgctac cgggggcgac gccgacgaaa 1320cccggttcgg caacgctgcc gttccctggt attcaagcgg acgtgctgga tttggacgga 1380aacgaaattc cggcgaacca ggggggatat ttggtcgtca aacatccctg gccgggcatg 1440atgcggacgg tttacggaga ctttaaccgt tttcgccgca gctattggga gcatattgct 1500ccgaaagacg gtcagtattt ctattttgcc ggagacggcg ctcgcaagga cgaggacggc 1560tatttctgga ttatgggtcg cgtggacgac gtgatcaacg tttcgggaca tcgcctcggg 1620acgatggaaa tcgagtcggc gttggtgtcg cacccgtcag tggcggaagc ggcggtggtc 1680gggaagccgg acgagattaa gggtgaaagc atcgtggcgt tcgtgatgtt ggaggaggac 1740tacgaggctg gcgacgactt ggataaggcg ttgaagcagc acgtggttga ggaaatcggc 1800gcgatcgccc gtccgggtga gattcgtttt tcagaagatt tgccgaaaa 184912616PRTGeitlerinema sp. 12Met Ser Glu Gln Thr Ile Glu Ser Ile Leu His Glu Gln Arg Thr Phe 1 5 10 15 Pro Pro Ala Ala Asp Phe Ala Ala Asn Ala His Ile Lys Ser Met Ala 20 25 30 Asp Tyr Lys Ala Leu Cys Asp Arg Ala Glu Lys Asp Pro Ala Gly Phe 35 40 45 Trp Ser Glu Leu Ala Glu Thr Glu Leu Asp Trp Phe Gln Lys Trp Glu 50 55 60 Asn Val Leu Asp Trp Gln Pro Pro Val Ala Lys Trp Phe Glu Gly Gly 65 70 75 80 Lys Leu Asn Val Ser Tyr Asn Cys Leu Asp Arg His Leu Thr Thr Trp 85 90 95 Arg Lys Asn Lys Ala Ala Leu Ile Trp Glu Gly Glu Pro Gly Asp Ser 100 105 110 Arg Thr Leu Thr Tyr Ala Gln Leu His Arg Glu Val Cys Gln Met Ala 115 120 125 Asn Val Ile Lys Gln Phe Gly Val Lys Lys Gly Asp Val Val Gly Ile 130 135 140 Tyr Met Pro Met Ile Pro Glu Ala Ala Ile Ala Met Leu Ala Cys Ala 145 150 155 160 Arg Ile Gly Ala Val His Ser Val Val Phe Gly Gly Phe Ser Ala Glu 165 170 175 Ala Leu Arg Asp Arg Val Asn Ala Ala Glu Ala Lys Leu Val Ile Thr 180 185 190 Ala Asp Gly Gly Phe Arg Lys Asp Lys Val Val Thr Leu Lys Asp Gln 195 200 205 Val Asp Lys Ala Leu Ala Asn Asp Ala Ala Pro Ser Val Glu Asn Val 210 215 220 Leu Val Val Arg Arg Ile Glu Lys Asp Thr His Met Glu Glu Gly Arg 225 230 235 240 Asp His Trp Trp His Glu Val Arg Gln Gly Ile Ser Ala His Cys Pro 245 250 255 Ala Glu Pro Met Asp Ser Glu Asp Ile Leu Phe Ile Leu Tyr Thr Ser 260 265 270 Gly Ser Thr Gly Lys Pro Lys Gly Val Val His Thr Thr Ala Gly Tyr 275 280 285 Asn Leu Tyr Ala His Val Thr Asn Lys Trp Thr Phe Asp Leu Lys Asp 290 295 300 Thr Asp Ile Phe Trp Cys Thr Ala Asp Val Gly Trp Ile Thr Gly His 305 310 315 320 Ser Tyr Ile Val Tyr Gly Pro Leu Ser Asn Gly Ala Thr Thr Val Met 325 330 335 Tyr Glu Gly Val Pro Arg Pro Ser Asn Pro Gly Cys Phe Trp Asp Val 340 345 350 Val Glu Lys Tyr Gly Val Thr Ile Phe Tyr Thr Ala Pro Thr Ala Ile 355 360 365 Arg Ala Phe Ile Lys Ala Gly Asp Lys His Pro Asn Ala Arg Asp Leu 370 375 380 Ser Ser Leu Arg Leu Leu Gly Thr Val Gly Glu Pro Ile Asn Pro Lys 385 390 395 400 Ala Trp Met Trp Tyr His Arg Val Ile Gly Gly Glu Arg Cys Pro Val 405 410 415 Val Asp Thr Trp Trp Gln Thr Glu Thr Gly Gly Phe Met Ile Thr Pro 420 425 430 Leu Pro Gly Ala Thr Pro Thr Lys Pro Gly Ser Ala Thr Leu Pro Phe 435 440 445 Pro Gly Ile Gln Ala Asp Val Leu Asp Leu Asp Gly Asn Glu Ile Pro 450 455 460 Ala Asn Gln Gly Gly Tyr Leu Val Val Lys His Pro Trp Pro Gly Met 465 470 475 480 Met Arg Thr Val Tyr Gly Asp Phe Asn Arg Phe Arg Arg Ser Tyr Trp 485 490 495 Glu His Ile Ala Pro Lys Asp Gly Gln Tyr Phe Tyr Phe Ala Gly Asp 500 505 510 Gly Ala Arg Lys Asp Glu Asp Gly Tyr Phe Trp Ile Met Gly Arg Val 515 520 525 Asp Asp Val Ile Asn Val Ser Gly His Arg Leu Gly Thr Met Glu Ile 530 535 540 Glu Ser Ala Leu Val Ser His Pro Ser Val Ala Glu Ala Ala Val Val 545 550 555 560 Gly Lys Pro Asp Glu Ile Lys Gly Glu Ser Ile Val Ala Phe Val Met 565 570 575 Leu Glu Glu Asp Tyr Glu Ala Gly Asp Asp Leu Asp Lys Ala Leu Lys 580 585 590 Gln His Val Val Glu Glu Ile Gly Ala Ile Ala Arg Pro Gly Glu Ile 595 600 605 Arg Phe Ser Glu Asp Leu Pro Lys 610 615 131410DNAGeitlerinema sp. 13atggaatcgg tgcaaactcc atcgccgcag actccatcgt cagtcgccga tcgcgtccga 60gcgcaacggg cgttcttcgc gacgggcaaa accaaagacg taaatttccg cctcgaacaa 120ctcaaacgcc tcaaacacgc gattctcgac taccgagacc gaatcgtcga agcagtcggg 180gctgacttgc gccgcccgga atttgaagcc tatttcgaga tcgcctccat cgccgaagtc 240aacaccgcga tcgcacgcct gaaatcttgg gcgaaaccca aacgggtatc cacctctctc 300gatcagtttc cgtctcgcgc ccgcattcac cccgaaccgt tgggcgtcgt gctaatcgtc 360gccccctgga actacccgtt tcaactgacg atgagtcccc tcgtcggcgc gatcgcggcc 420ggaaattgcg ccgttctcaa accctcggaa attgcccccc acaccgcagc cgtcgtcagt 480gacttgattc gctcgacctt tccccctgaa tacgtcaccg ccatcgaagg cggcgtcgaa 540accagtcaat ccctcctcga acagaaattc gacaaaatct tttttaccgg gggaacccgc 600atcggccaga tcgtcatgga agcggcggcg aaacacctca cccccgttac cctggaactc 660ggcggaaaaa gcccctgtat cgtcgatgcg gacgtgaaac tcgacgttgc tgtcaaacgc 720atcgtttggg gaaagtttat caacgccgga caaacctgtg tcgcgccgga ttacctgctc 780gtcgatcgcc gcgttaaacc ccgactgctc gaagcggtgc gccagcaagt ccgcgagttt 840tttggcgacg atcccgccaa aagtgccgat ttctgtcgca tggtaagcga tcgccatttc 900gatcgcgtcg cctcgttact agaaaatcgg ggaaatgctg agattgtcgt cggcggacag 960tgcgatcgca gcgatcgcta catcgccccc accgtcctcg ataacgtatc ctggaacgat 1020ccggtgatgc aagacgaaat tttcggcccg attctgcccg ttttagagta cgacagtctc 1080gacgacgcca tcgatcgcgt cgcttcccgt cccaaacccc tcgccctcta cgtcttttct 1140aacaacaaac ccttccaaaa ccgcgtcttg cgcgagactt cctccggcgg agcctgcgtc 1200aacgatactg tcatgcacct ggccgtttcc gatctcccct ttggcggcgt cggcgacagc 1260ggaatgggaa gctatcacgg aaaagccagt ttcgatacct tctcccattt caaaagcgtc 1320ctcaacaaag gactttggtt cgatctcaac tggcgttacg cgccctatca tcaatggcaa 1380ctcagccttc tcaaacgcat catcggttga 141014469PRTGeitlerinema sp. 14Met Glu Ser Val Gln Thr Pro Ser Pro Gln Thr Pro Ser Ser Val Ala 1 5 10 15 Asp Arg Val Arg Ala Gln Arg Ala Phe Phe Ala Thr Gly Lys Thr Lys 20 25 30 Asp Val Asn Phe Arg Leu Glu Gln Leu Lys Arg Leu Lys His Ala Ile 35 40 45 Leu Asp Tyr Arg Asp Arg Ile Val Glu Ala Val Gly Ala Asp Leu Arg 50 55 60 Arg Pro Glu Phe Glu Ala Tyr Phe Glu Ile Ala Ser Ile Ala Glu Val 65 70 75 80 Asn Thr Ala Ile Ala Arg Leu Lys Ser Trp Ala Lys Pro Lys Arg Val 85 90 95 Ser Thr Ser Leu Asp Gln Phe Pro Ser Arg Ala Arg Ile His Pro Glu 100 105 110 Pro Leu Gly Val Val Leu Ile Val Ala Pro Trp Asn Tyr Pro Phe Gln 115 120 125 Leu Thr Met Ser Pro Leu Val Gly Ala Ile Ala Ala Gly Asn Cys Ala 130 135 140 Val Leu Lys Pro Ser Glu Ile Ala Pro His Thr Ala Ala Val Val Ser 145 150 155 160 Asp Leu Ile Arg Ser Thr Phe Pro Pro Glu Tyr Val Thr Ala Ile Glu 165 170 175 Gly Gly Val Glu Thr Ser Gln Ser Leu Leu Glu Gln Lys Phe Asp Lys 180 185 190 Ile Phe Phe Thr Gly Gly Thr Arg Ile Gly Gln Ile Val Met Glu Ala 195 200 205 Ala Ala Lys His Leu Thr Pro Val Thr Leu Glu Leu Gly Gly Lys Ser 210 215 220 Pro Cys Ile Val Asp Ala Asp Val Lys Leu Asp Val Ala Val Lys Arg 225 230 235 240 Ile Val Trp Gly Lys Phe Ile Asn Ala Gly Gln Thr Cys Val Ala Pro 245 250 255 Asp Tyr Leu Leu Val Asp Arg Arg Val Lys Pro Arg Leu Leu Glu Ala 260 265 270 Val Arg Gln Gln Val Arg Glu Phe Phe Gly Asp Asp Pro Ala Lys Ser 275 280 285 Ala Asp Phe Cys Arg Met Val Ser Asp Arg His Phe Asp Arg Val Ala 290 295 300 Ser Leu Leu Glu Asn Arg Gly Asn Ala Glu Ile Val Val Gly Gly Gln 305 310 315 320 Cys Asp Arg Ser Asp Arg Tyr Ile Ala Pro Thr Val Leu Asp Asn Val 325 330 335 Ser Trp Asn Asp Pro Val Met Gln Asp Glu Ile Phe Gly Pro Ile Leu 340 345 350 Pro Val Leu Glu Tyr Asp Ser Leu Asp Asp Ala Ile Asp Arg Val Ala 355 360 365 Ser Arg Pro Lys Pro Leu Ala Leu Tyr Val Phe Ser Asn Asn Lys Pro 370 375 380 Phe Gln Asn Arg Val Leu Arg Glu Thr Ser Ser Gly Gly Ala Cys Val 385 390 395 400 Asn Asp Thr Val Met His Leu Ala Val Ser Asp Leu Pro Phe Gly Gly 405 410 415 Val Gly Asp Ser Gly Met Gly Ser Tyr His Gly Lys Ala Ser Phe Asp 420 425 430 Thr Phe Ser His Phe Lys Ser Val Leu Asn Lys Gly Leu Trp Phe Asp 435 440 445 Leu Asn Trp Arg Tyr Ala Pro Tyr His Gln Trp Gln Leu Ser Leu Leu 450 455 460 Lys Arg Ile Ile Gly 465 151218DNAGeitlerinema sp. 15atgaagattc tcgtactgaa tgctggctcg agttctcaga aaagctgttt gtacgacgtt 60cccggtgcgg ggtttcccga tacgccccaa gaaccgattt gggaagcaac catcgattgg 120ggcgtgggga cggaatacgg actgttaact gtcgaagcca acgagacgaa gcagaaaagc 180gaactctcga tttacgctcg ggcccacggt ttgggggaaa tgctggatac gctggtgcag 240ggcgaaacga aggttctcga ggacttgtcg gaaattgcga tcgtcggtca tcgggtggtt 300cacgggggaa cggagtattc ggacgctacg tatattacgc cagcggtgaa acaagccatt 360gaagatttga ttcccttagc ccccaaccac aaccccgctc atttagaaga aatcctcgct 420gtcgaggaag tgttgggaga cgtaccacaa gtggcggtat tcgatacggc gtttcacagt 480caaatgccga catcggtggc ggcgtatccg attccctatc gctggtttga aaagggagtg

540cgtcggtatg ggttccacgg tatcagccat cgctactgtg ccgaacgtgc ggcggaactc 600ctcgaggagc cgttggaatc gctgcgaatc gtaacttgtc atttgggaca tggctgttct 660ctggcggcgg ttcgcgacgg aatgagtgtg aatacgacga tgggtttcac gccgttggaa 720gggttgatga tggggagtcg tagcggttcg atcgatccgg cgattcccat gtatttgatg 780cgcgaggaag ggttcgattt cgagggcgtg gataagatgc tgaataagga atcgggactc 840aaaggcgttt ctggtgagtc gggggatatg cgatcgatcc tcaaggcgat gggagagggg 900agcgatcgcg cagagttggc gttcgagatg tacgtctctc gattgcagag tgcgatcgcc 960tcgatgattc cccagttagg ggggttagat gttttggcat ttacggcggg tgtcggcgag 1020aattccgccg acgtgcgagc ggcaacttgt gcggggttag attttttggg cttaaaactc 1080gattcgcgcc aaaacgccgc gtctcccaag gatgcggata tcgcgtcaat ggattcgacg 1140gtgcgggtgt tggtgattcg cgctcaggag gattgggcga tcgcggggga atgttggaag 1200ttggtaaagg taggttag 121816405PRTGeitlerinema sp. 16Met Lys Ile Leu Val Leu Asn Ala Gly Ser Ser Ser Gln Lys Ser Cys 1 5 10 15 Leu Tyr Asp Val Pro Gly Ala Gly Phe Pro Asp Thr Pro Gln Glu Pro 20 25 30 Ile Trp Glu Ala Thr Ile Asp Trp Gly Val Gly Thr Glu Tyr Gly Leu 35 40 45 Leu Thr Val Glu Ala Asn Glu Thr Lys Gln Lys Ser Glu Leu Ser Ile 50 55 60 Tyr Ala Arg Ala His Gly Leu Gly Glu Met Leu Asp Thr Leu Val Gln 65 70 75 80 Gly Glu Thr Lys Val Leu Glu Asp Leu Ser Glu Ile Ala Ile Val Gly 85 90 95 His Arg Val Val His Gly Gly Thr Glu Tyr Ser Asp Ala Thr Tyr Ile 100 105 110 Thr Pro Ala Val Lys Gln Ala Ile Glu Asp Leu Ile Pro Leu Ala Pro 115 120 125 Asn His Asn Pro Ala His Leu Glu Glu Ile Leu Ala Val Glu Glu Val 130 135 140 Leu Gly Asp Val Pro Gln Val Ala Val Phe Asp Thr Ala Phe His Ser 145 150 155 160 Gln Met Pro Thr Ser Val Ala Ala Tyr Pro Ile Pro Tyr Arg Trp Phe 165 170 175 Glu Lys Gly Val Arg Arg Tyr Gly Phe His Gly Ile Ser His Arg Tyr 180 185 190 Cys Ala Glu Arg Ala Ala Glu Leu Leu Glu Glu Pro Leu Glu Ser Leu 195 200 205 Arg Ile Val Thr Cys His Leu Gly His Gly Cys Ser Leu Ala Ala Val 210 215 220 Arg Asp Gly Met Ser Val Asn Thr Thr Met Gly Phe Thr Pro Leu Glu 225 230 235 240 Gly Leu Met Met Gly Ser Arg Ser Gly Ser Ile Asp Pro Ala Ile Pro 245 250 255 Met Tyr Leu Met Arg Glu Glu Gly Phe Asp Phe Glu Gly Val Asp Lys 260 265 270 Met Leu Asn Lys Glu Ser Gly Leu Lys Gly Val Ser Gly Glu Ser Gly 275 280 285 Asp Met Arg Ser Ile Leu Lys Ala Met Gly Glu Gly Ser Asp Arg Ala 290 295 300 Glu Leu Ala Phe Glu Met Tyr Val Ser Arg Leu Gln Ser Ala Ile Ala 305 310 315 320 Ser Met Ile Pro Gln Leu Gly Gly Leu Asp Val Leu Ala Phe Thr Ala 325 330 335 Gly Val Gly Glu Asn Ser Ala Asp Val Arg Ala Ala Thr Cys Ala Gly 340 345 350 Leu Asp Phe Leu Gly Leu Lys Leu Asp Ser Arg Gln Asn Ala Ala Ser 355 360 365 Pro Lys Asp Ala Asp Ile Ala Ser Met Asp Ser Thr Val Arg Val Leu 370 375 380 Val Ile Arg Ala Gln Glu Asp Trp Ala Ile Ala Gly Glu Cys Trp Lys 385 390 395 400 Leu Val Lys Val Gly 405

Claims

1. A biofilm photobioreactor system comprising: a) a flexible film defining a photobioreactor enclosure; b) channels defined by at least one partition disposed in the photobioreactor enclosure, wherein the channels are in fluid communication; c) a support substrate suitable for the formation of a biofilm thereon by a microorganism, wherein the support substrate is disposed in the channels and is fixed to the flexible film, the at least one partition or a combination of the flexible film and the at least one partition; d) a photosynthetic, autofermentative microorganism forming a biofilm on the support substrate, wherein the photosynthetic, fermentative microorganism makes at least one metabolic intermediate compound during a photosynthesis phase and converts the at least one metabolic intermediate compound to at least one chemical product during an autofermentation phase; e) at least one port formed in the flexible film that is in fluid communication with the photobioreactor enclosure; f) at least one gas exhaust vent formed in the flexible film that is in fluid communication with the photobioreactor enclosure; g) gas comprising carbon dioxide that flows through the photobioreactor enclosure intermittently during the photosynthesis phase, wherein the gas enters the photobioreactor enclosure through the at least one port, contacts the biofilm and leaves the photobioreactor enclosure through the at least one gas exhaust vent; h) a first liquid that at least partially fills the photobioreactor enclosure intermittently during the photosynthesis phase, wherein the first liquid enters the photobioreactor enclosure through the at least one port, contacts the biofilm and leaves the photobioreactor enclosure through the at least one port; and i) a second liquid that at least partially fills the photobioreactor enclosure during the autofermentation phase, wherein the second liquid enters the photobioreactor enclosure through the at least one port, contacts the biofilm and leaves the photobioreactor enclosure through the at least one port and wherein the at least one chemical product made by the biofilm during the autofermentation phase enters the second liquid; wherein the photobioreactor enclosure has a flat panel shape and at least one portion of the flexible film is translucent.

2. The biofilm photobioreactor system of claim 1 wherein the at least one metabolic intermediate is selected from the group consisting of carbohydrates and osmoprotectants.

3. The biofilm photobioreactor system of claim 2 wherein the carbohydrate is glycogen.

4. The biofilm photobioreactor system of claim 2 wherein the osmoprotectant is selected from the group consisting of trehalose and glucosylglycerol.

5. The biofilm photobioreactor system of claim 1 wherein the at least one chemical product is selected from the group consisting of biofuels and feedstocks.

6. The biofilm photobioreactor system of claim 5 wherein the biofuel is selected from the group consisting of ethanol, hydrogen, propanol and butanol.

7. The biofilm photobioreactor system of claim 5 wherein the feedstock is selected from the group consisting of acetate, lactate and formate.

8. The biofilm photobioreactor system of claim 1 further comprising a mounting system that provides external support for at least one biofilm photobioreactor.

9. The biofilm photobioreactor system of claim 8 wherein the mounting system is adapted to adjust the position of the at least one biofilm photobioreactor between vertical and horizontal.

10. The biofilm photobioreactor system of claim 1 wherein the orientation of the biofilm photobioreactor is substantially vertical.

11. The biofilm photobioreactor system of claim 1 wherein the photosynthetic, fermentative microorganism is selected from the group consisting of Geitlerinema, Lyngbya, Chroococcidiopsis, Calothrix, Cyanothece, Oscillatoria, Gloeothece, Microcoleus, Microcystis, Nostoc, Anabaena and Spirulina species.

12. The biofilm photobioreactor system of claim 1 wherein the support substrate comprises at least one material selected from the group consisting of films, filters, fabrics, foams and felts of polyesters, polyolefins, polyurethanes, polyamides, polyimides, polycarbonates, polydienes and polyacrylics.

13. The biofilm photobioreactor system of claim 1 wherein the first liquid is a freshwater medium.

14. The biofilm photobioreactor system of claim 1 wherein the second liquid is a freshwater medium that is substantially depleted of electron acceptors.

15. The biofilm photobioreactor system of claim 1 wherein the second liquid is a freshwater medium that is sparged with nitrogen gas.

16. The biofilm photobioreactor system of claim 1 wherein the flexible film comprises at least one material selected from the group consisting of polyolefins, polyesters and vinyl copolymers thereof.

17. The biofilm photobioreactor system of claim 1 wherein the at least one port is positioned near the bottom edge of the photobioreactor enclosure when the biofilm photobioreactor is vertical.

18. The biofilm photobioreactor system of claim 1 wherein the at least one gas exhaust vent is positioned near the top edge of the photobioreactor enclosure when the biofilm photobioreactor is vertical.

19. The biofilm photobioreactor system of claim 1 wherein the gas is air.

20. A flat panel biofilm photobioreactor system comprising: a) a plurality of channels in fluid communication that are adapted to admit light; b) inlets and outlets adapted to admit flow of a gas comprising carbon dioxide and flow of a liquid into and out of the channels; c) a formation surface disposed in the channels; and d) a photosynthetic, autofermentative microorganism that makes a chemical product through autofermentation and forms a biofilm on the formation surface; wherein the channels are formed in an enclosure enclosed by a flexible film, the formation surface is fixed to the flexible film, the gas comprising carbon dioxide and the liquid contact the biofilm and the chemical product enters the liquid.

21. A method of making at least one chemical product comprising the steps of: a) exposing a biofilm comprising a photosynthetic, autofermentative microorganism to light, wherein the biofilm is disposed on a support substrate that is disposed in channels of a biofilm photobioreactor and wherein the support substrate is fixed to inner surfaces of the biofilm photobioreactor, b) alternately flowing a gas comprising carbon dioxide through the channels and at least partially filling the channels with a first liquid, wherein the gas comprising carbon dioxide and the first liquid contact the biofilm and the photosynthetic, autofermentative microorganism makes at least one metabolic intermediate compound by photosynthesis; c) depriving the biofilm of light; and d) at least partially filling the channels of the biofilm photobioreactor with a second liquid, wherein the second liquid contacts the biofilm, the photosynthetic, autofermentative microorganism converts the at least one metabolic intermediate compound into the at least one chemical product and the at least one chemical product enters the second liquid.

22. The method of claim 21 further comprising the step of exerting a pulling force perpendicular to the longitudinal axes of the chambers before at least partially filling the chambers of the biofilm photobioreactor with a second liquid.

23. The method of claim 21 further comprising the step of extracting the second liquid containing the at least one chemical product from the biofilm photobioreactor.

24. The method of claim 21 further comprising the step of separating the at least one chemical product from the second liquid.

25. The method of claim 21 wherein the gas comprising carbon dioxide flows through the chambers for at least one time period of from about 5 minutes to about 4 hours.

26. The method of claim 21 wherein the chambers are at least partially filled with the first liquid for at least one time period of from about 10 seconds to about 20 minutes.

27. The method of claim 21 wherein the chambers are at least partially filled with the second liquid for a time period of from about 1 hour to about 18 hours.

28. The method of claim 21 wherein the second liquid is substantially depleted of electron acceptors.

29. The method of claim 21 further comprising the step of sparging the second liquid with nitrogen gas prior to at least partially filling the chambers of the biofilm photobioreactor with the second liquid.

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