Use of Cellulosic Materials for Cultivation of Microorganisms

• US 20090011480A1
• Filed: 06/02/2008
• Published: 01/08/2009
• Est. Priority Date: 06/01/2007
• Status: Active Grant

• Current Assignee: Corbion Biotech, Inc. (Corbion NV)
• Original Assignee: Solazyme Inc.
• Inventors: Franklin, Scott, Day, Anthony G., Dillon, Harrison F., Trimbur, Donald E., Im, Chung-Soon, Coragliotti, Anna
• Granted Patent: US 8,790,914 B2
• Time in Patent Office: Days
• US Class Current: 435/134
• CPC Class Codes: C07H 21/04 with deoxyribosyl as saccharide radical
• CPC Class Codes: C10L 1/026 for compression ignition
• CPC Class Codes: C10L 1/06 for spark ignition
• CPC Class Codes: C10L 2200/0484 Vegetable or animal oils
• CPC Class Codes: C10L 2270/026 for diesel engines, e.g. automobiles, stationary, marine
• CPC Class Codes: C10L 2270/04 for turbines, planes, power generation
• CPC Class Codes: C11C 3/00 Fats, oils, or fatty acids by chemical modification of fats, oils, or fatty acids obtained therefrom sulfonated fats or oils C07C309/62; factice C08H; drying oils C09F
• CPC Class Codes: C12N 1/12 Unicellular algae; Culture media therefor as new plants A01H13/00
• CPC Class Codes: C12N 1/32 Processes using, or culture media containing, lower alkanols, i.e. C1 to C6
• CPC Class Codes: C12N 15/79 Vectors or expression systems specially adapted for eukaryotic hosts
• CPC Class Codes: C12N 9/0006 acting on CH-OH groups as donors (1.1)
• CPC Class Codes: C12N 9/0008 acting on the aldehyde or oxo group of donors (1.2)
• CPC Class Codes: C12N 9/1205 Phosphotransferases with an alcohol group as acceptor (2.7.1), e.g. protein kinases
• CPC Class Codes: C12N 9/14 Hydrolases (3)
• CPC Class Codes: C12N 9/16 acting on ester bonds (3.1)
• CPC Class Codes: C12N 9/20 Triglyceride splitting, e.g. by means of lipase
• CPC Class Codes: C12N 9/2408 acting on alpha -1,4-glucosidic bonds
• CPC Class Codes: C12N 9/2431 Beta-fructofuranosidase (3.2.1.26), i.e. invertase
• CPC Class Codes: C12N 9/88 Lyases (4.)
• CPC Class Codes: C12P 21/06 produced by the hydrolysis of a peptide bond, e.g. hydrolysate products preparing foodstuffs by protein hydrolysis A23J3/00
• CPC Class Codes: C12P 5/02 : acyclic C12P5/007 takes precedence
• CPC Class Codes: C12P 7/6409 : Fatty acids
• CPC Class Codes: C12P 7/6458 : by transesterification, e.g. interesterification, ester interchange, alcoholysis or acidolysis
• CPC Class Codes: C12P 7/6463 : obtained from glyceride producing microorganisms, e.g. single cell oil
• CPC Class Codes: C12P 7/649 : Biodiesel, i.e. fatty acid alkyl esters
• CPC Class Codes: C12Y 302/01026 : Beta-fructofuranosidase (3.2.1.26), i.e. invertase
• CPC Class Codes: Y02E 50/10 : Biofuels, e.g. bio-diesel
• CPC Class Codes: Y02E 50/30 : Fuel from waste, e.g. synthetic alcohol or diesel
• CPC Class Codes: Y02P 20/52 : using catalysts, e.g. selective catalysts
• CPC Class Codes: Y02T 50/678 : Aviation using fuels of non-fossil origin

Abstract

The invention provides methods of cultivating oil-bearing microbes using cellulosic material. Also provided are microorganisms that manufacture non-alcohol-based fuels and fuel feedstocks through a process of converting cellulosic materials into oils. Also provided are compositions comprising depolymerized cellulosic materials and oil-bearing microbes. Some methods of microbial fermentation are provided that comprise combining depolymerized cellulosic materials with other non-cellulosic feedstocks to enhance the economics of renewable fuel manufacturing. Particular advantages of the processes provided herein include production of oils rather than alcohols through cellulosic processes. Additional advantages include methods for manufacturing high nutrition oils from non-edible feedstocks such as wood chips, switchgrass, and bagasse.

Claims

1. A method of fermenting a microorganism comprising providing a mixture comprising glucose and xylose as an energy source to the microorganism.

2. The method of claim 1, wherein the mixture further comprises lignin.

3. The method of claim 1, wherein the mixture further comprises at least one species of furfural.

4. The method of claim 1, wherein the mixture is depolymerized cellulosic material.

5. The method of claim 1, wherein the microorganism is a microalgae.

6. The method of claim 5, wherein the microalgae is selected from Table 1.

7. The method of claim 6, wherein the microalgae is a species of the genus Chlorella.

8. The method of claim 7, wherein the microalgae is selected from the group consisting of Chlorella anitrata, Chlorella antarctica, Chlorella aureoviridis, Chlorella candida, Chlorella capsulata, Chlorella desiccata, Chlorella ellipsoidea, Chlorella emersonii, Chlorella fusca, Chlorella fusca var. vacuolata, Chlorella glucotropha, Chlorella infusionum, Chlorella infusionum var. Actophila, Chlorella infusionum var. Auxenophila, Chlorella kessleri, Chlorella luteoviridis, Chlorella luteoviridis var. aureoviridis, Chlorella luteoviridis var. Lutescens, Chlorella miniata, Chlorella minutissima, Chlorella mutabilis, Chlorella nocturna, Chlorella parva, Chlorella photophila, Chlorella pringsheimii, Chlorella protothecoides, Chlorella pyrenoidosa, Chlorella regularis, Chlorella regularis var. minima, Chlorella regularis var. umbricata, Chlorella reisiglii, Chlorella saccharophila, Chlorella saccharophila var. ellipsoidea, Chlorella salina, Chlorella simplex, Chlorella sorokiniana, Chlorella sp., Chlorella sphaerica, Chlorella stigmatophora, Chlorella vanniellii, Chlorella vulgaris, Chlorella vulgaris, Chlorella vulgarisf tertia, Chlorella vulgaris var. airidis, Chlorella vulgaris var. vulgaris, Chlorella vulgaris var. vulgarisf tertia, Chlorella vulgaris var. vulgarisf viridis, Chlorella xanthella, and Chlorella zofingiensis.

9. The method of claim 6, wherein the microalgae is selected from the group consisting of Bracteococcus minor, Chlorella ellipsoidea, Chlorella kessleri, Chlorella luteoviridis, Bracteococcus medionucleatus, Chlorella minutissima, Chlorella ovalis, Chlorella protothecoides, Chlorella saccharophila, Chlorella sorokiniana, Chlorella sp., Chlorella vulgaris, Parachlorella kessleri, Prototheca moriformis, and Pseudochlorella aquatica.

10. The method of claim 1, wherein the microorganism is an oleaginous yeast.

11. The method of claim 10, wherein the oleaginous yeast is selected from the group consisting of Cryptococcus curvatus, Cryptococcus terricolus, Candida sp., Lipomyces starkeyi, Lipomyces lipofer, Endomycopsis vernalis, Rhodotorula glutinis, Rhodotorula gracilis, and Yarrowia lipolytica.

12. The method of claim 1, wherein the microorganism is a fungus.

13. The method of claim 12, wherein the fungus is selected from the group consisting of a species of the genus Mortierella, Mortierrla vinacea, Mortierella alpine, Pythium debaryanum, Mucor circinelloides, Aspergillus ochraceus, Aspergillus terreus, Pennicillium iilacinum, a species of the genus Hensenulo, a species of the genus Chaetomium, a species of the genus Cladosporium, a species of the genus Malbranchea, a species of the genus Rhizopus, and a species of the genus Pythium.

14. The method of claim 1, wherein the mixture further comprises at least one sucrose utilization enzyme.

15. The method of claim 14, wherein the microorganism is of the genus Chlorella.

16. The method of claim 1, wherein the microorganism has been genetically engineered to express an exogenous gene encoding at least one lipid modification enzyme, hydrocarbon modification enzyme, or sucrose utilization enzyme.

17. The method of claim 14, wherein the mixture comprises a sucrose invertase.

59. The method of claim 1, wherein the mixture further comprises sucrose.

60. The method of claim 4, wherein the depolymerized cellulosic material is one or more materials selected from the group consisting of sugarcane bagasse, rice hulls, corn stover, wheat straw, rice straw, sugar beet pulp, citrus pulp, citrus peels, hardwood thinnings, softwood thinnings, wood chips, sawdust, pulp mill waste, urban paper waste, grass clippings, switchgrass, hybrid poplar wood, miscanthus, fiber cane, and fiber sorghum.

18. A method of culturing a microalgae comprising culturing the microalgae in a culture medium including a feedstock comprising at least one carbon substrate selected from the group consisting of a depolymerized cellulosic material, a 5-carbon sugar, a 6-carbon sugar, and acetate.

19. The method of claim 18, wherein the carbon substrate is glucose and the microalgae is of a genus selected from the group consisting of Chlorella, Parachlorella, Pseudochlorella, Bracteococcus, Prototheca and Scenedesmus.

20. The method of claim 18, wherein the carbon substrate is xylose and the microalgae is of a genus selected from the group consisting of Chlorella, Pseudochlorella, and Prototheca.

21. The method of claim 18, wherein the carbon substrate is sucrose and the microalgae is of a genus selected from the group consisting of Chlorella, and Bracteococcus.

22. The method of claim 18, wherein the carbon substrate is fructose and the microalgae is of a genus selected from the group consisting of Chlorella, Parachlorella, Prototheca, and Scenedesmus.

23. The method of claim 18, wherein the carbon substrate is arabinose and the microalgae is Chlorella sp.

24. The method of claim 18, wherein the carbon substrate is mannose and the microalgae is of a genus selected from the group consisting of Chlorella, Parachlorella, Bracteococcus, Prototheca, and Scenedesmus.

25. The method of claim 18, wherein the carbon substrate is galactose and the microalgae is of a genus selected from the group consisting of Bracteococcus, Parachlorella, Chlorella, Pseudochlorella, Bracteococcus, and Prototheca.

26. The method of claim 18, wherein the carbon substrate is acetate and the microalgae is of a genus selected from the group consisting of Chlorella, Parachlorella, and Prototheca.

27. The method of claim 18, wherein the culture medium further includes at least one sucrose utilization enzyme.

28. The method of claim 27, wherein the microalgae is of the genus Chlorella.

29. The method of claim 28, wherein the microalgae has been genetically engineered to express an exogenous gene encoding at least one lipid modification enzyme, hydrocarbon modification enzyme, or sucrose utilization enzyme.

30. The method of claim 27, wherein the culture medium includes a sucrose invertase.

31. The method of claim 18, wherein the microalgae is selected from Table 1.

61. The method according to claim 18, wherein the depolymerized cellulosic material is one or more materials selected from the group consisting of sugarcane bagasse, rice hulls, corn stover, wheat straw, rice straw, sugar beet pulp, citrus pulp, citrus peels, hardwood thinnings, softwood thinnings, wood chips, sawdust, pulp mill waste, urban paper waste, grass clippings, switchgrass, hybrid poplar wood, miscanthus, fiber cane, and fiber sorghum.

62. The method according to claim 18, wherein the feedstock comprises depolymerized cellulosic material and acetate.

32. A method of culturing microalgae comprising placing a plurality of microalgae cells in the presence of depolymerized cellulosic material.

33. The method of claim 32, wherein the microalgae are cultured in the presence of an additional fixed carbon source selected from the group consisting of glycerol, sucrose, glucose, arabinose, galactose, xylose, fructose, arabinose, mannose, acetate, and any combination of the foregoing.

34. The method of claim 32, wherein the microalgae is selected from a species of the genus Bracteococcus, a species of the genus Chlorella, a species of the genus Parachlorella, a species of the genus Prototheca, or a species of the genus Pseudochlorella.

35. The method of claim 34, wherein the microalgae is selected from Bracteococcus minor, Chlorella ellipsoidea, Chlorella kessleri, Chlorella luteoviridis, Bracteococcus medionucleatus, Chlorella minutissima, Chlorella ovalis, Chlorella protothecoides, Chlorella saccharophila, Chlorella sorokiniana, Chlorella sp., Chlorella vulgaris, Parachlorella kessleri, Prototheca moriformis, and Pseudochlorella aquatica.

36. The method of claim 32, wherein the microalgae are cultured in the presence of at least one sucrose utilization enzyme.

37. The method of claim 36, wherein the microalgae is of the genus Chlorella.

38. The method of claim 32, wherein the microalgae has been genetically engineered to express an exogenous gene encoding at least one lipid modification enzyme, hydrocarbon modification enzyme, or sucrose utilization enzyme.

39. The method of claim 36, wherein the at least one sucrose utilization enzyme is a sucrose invertase.

40. The method of claim 38, wherein the at least one lipid modification enzyme is selected from a stearoyl-ACP desaturase, a glycerolipid desaturase, a pyruvate dehydrogenase, an acetyl-CoA carboxylase, and a glycerol-3 phosphate acyltransferase.

41. The method of claim 38, wherein the at least one hydrocarbon modification enzyme is selected from a fatty acyl-ACP thioesterase, a fatty acyl-CoA reductase, a fatty aldehyde reductase, a fatty acyl-CoA/aldehyde reductase, a fatty aldehyde decarbonylase, and an acyl carrier protein.

63. The method according to claim 32, wherein the depolymerized cellulosic material is one or more materials selected from the group consisting of sugarcane bagasse, rice hulls, corn stover, wheat straw, rice straw, sugar beet pulp, citrus pulp, citrus peels, hardwood thinnings, softwood thinnings, wood chips, sawdust, pulp mill waste, urban paper waste, grass clippings, switchgrass, hybrid poplar wood, miscanthus, fiber cane, and fiber sorghum.

42. A method of culturing a lipid-producing microbe, the method comprising culturing the microbe in the presence of acetate and the absence of a fixed nitrogen source.

43. The method of claim 42, wherein the microbe is cultured in the presence of a sufficient amount of acetate to increase microbial lipid yield over microbial lipid yield in the absence of acetate, wherein culture conditions are otherwise the same between the two cultures.

44. A microbial culture containing:

(a) a population of microorganisms; and

(b) a culture medium comprising glucose, xylose, and a molecule selected from the group consisting of lignin and a species of furfural.

45. The microbial culture of claim 44, wherein the microorganisms are microalgae.

46. The microbial culture of claim 45, wherein the microorganisms are selected from the group consisting of the microalgae listed in Table 1.

47. The microbial culture of claim 46, wherein the microorganisms are a species of the genus Chlorella.

48. The microbial culture of claim 47, wherein the microorganisms are selected from the group consisting of Chlorella anitrata, Chlorella antarctica, Chlorella aureoviridis, Chlorella candida, Chlorella capsulata, Chlorella desiccata, Chlorella ellipsoidea, Chlorella emersonii, Chlorella fusca, Chlorella fusca var. vacuolata, Chlorella glucotropha, Chlorella infusionum, Chlorella infusionum var. Actophila, Chlorella infusionum var. Auxenophila, Chlorella kessleri, Chlorella luteoviridis, Chlorella luteoviridis var. aureoviridis, Chlorella luteoviridis var. Lutescens, Chlorella miniata, Chlorella minutissima, Chlorella mutabilis, Chlorella nocturna, Chlorella parva, Chlorella photophila, Chlorella pringsheimii, Chlorella protothecoides, Chlorella pyrenoidosa, Chlorella regularis, Chlorella regularis var. minima, Chlorella regularis var. umbricata, Chlorella reisiglii, Chlorella saccharophila, Chlorella saccharophila var. ellipsoidea, Chlorella salina, Chlorella simplex, Chlorella sorokiniana, Chlorella sp., Chlorella sphaerica, Chlorella stigmatophora, Chlorella vanniellii, Chlorella vulgaris, Chlorella vulgaris, Chlorella vulgarisf tertia, Chlorella vulgaris var. airidis, Chlorella vulgaris var. vulgaris, Chlorella vulgaris var. vulgarisf tertia, Chlorella vulgaris var. vulgarisf viridis, Chlorella xanthella, and Chlorella zofingiensis.

49. The microbial culture of claim 44, wherein the microorganisms are selected from Bracteococcus minor, Chlorella ellipsoidea, Chlorella kessleri, Chlorella luteoviridis, Bracteococcus medionucleatus, Chlorella minutissima, Chlorella ovalis, Chlorella protothecoides, Chlorella saccharophila, Chlorella sorokiniana, Chlorella sp., Chlorella vulgaris, Parachlorella kessleri, Prototheca moriformis, and Pseudochlorella aquatica.

50. The microbial culture of claim 44, wherein the microorganisms are oleaginous yeast.

51. The microbial culture of claim 50, wherein the oleaginous yeast are selected from the group consisting of Cryptococcus curvatus, Cryptococcus terricolus, Candida sp., Lipomyces starkeyi, Lipomyces lipofer, Endomycopsis vernalis, Rhodotorula glutinis, Rhodotorula gracilis, and Yarrowia lipolytica.

52. The microbial culture of claim 44, wherein the microorganisms are fungi.

53. The microbial culture of claim 52, wherein the fungi are selected from the group consisting of a species of the genus Mortierella, Mortierrla vinacea, Mortierella alpine, Pythium debaryanum, Mucor circinelloides, Aspergillus ochraceus, Aspergillus terreus, Pennicillium iilacinum, a species of the genus Hensenulo, a species of the genus Chaetomium, a species of the genus Cladosporium, a species of the genus Malbranchea, a species of the genus Rhizopus, and a species of the genus Pythium.

54. The microbial culture of claim 44, wherein the microorganisms contain at least one exogenous sucrose utilization gene.

55. The microbial culture of claim 54, wherein the sucrose utilization gene encodes a sucrose transporter, a sucrose invertase, a hexokinase, a glucokinase, or a fructokinase.

56. The microbial culture of claim 44, wherein the microorganisms contain at least one exogenous gene encoding a lipid pathway enzyme.

57. The microbial culture of claim 56, wherein the lipid pathway enzyme is selected from the group consisting of a stearoyl-ACP desaturase, a glycerolipid desaturase, a pyruvate dehydrogenase, an acetyl-CoA carboxylase, an acyl carrier protein, and a glycerol-3 phosphate acyltransferase.

64. The method of claim 44, wherein the glucose, xylose, and a molecule selected from the group consisting of lignin and a species of furfural are derived from one or more depolymerized cellulosic feedstocks selected from the group consisting of sugarcane bagasse, rice hulls, corn stover, wheat straw, rice straw, sugar beet pulp, citrus pulp, citrus peels, hardwood thinnings, softwood thinnings, wood chips, sawdust, pulp mill waste, urban paper waste, grass clippings, switchgrass, hybrid poplar wood, miscanthus, fiber cane, and fiber sorghum.

58. A method of cultivating microalgae, comprising:

(a) providing a microalgae cell capable of performing heterotrophic growth;

(b) placing the microalgae cell in growth media, wherein the growth media comprises depolymerized cellulosic material; and

(c) incubating the microalgae for a period of time sufficient to allow the cell to grow.

65. The method of claim 58, wherein the depolymerized cellulosic material is selected from the group consisting of sugarcane bagasse, rice hulls, corn stover, wheat straw, rice straw, sugar beet pulp, citrus pulp, citrus peels, hardwood thinnings, softwood thinnings, wood chips, sawdust, pulp mill waste, urban paper waste, grass clippings, switchgrass, hybrid poplar wood, miscanthus, fiber cane, and fiber sorghum.

66. The method of claim 58, wherein the microalgae are selected from the group consisting of Chlorella anitrata, Chlorella antarctica, Chlorella aureoviridis, Chlorella candida, Chlorella capsulata, Chlorella desiccata, Chlorella ellipsoidea, Chlorella emersonii, Chlorella fusca, Chlorella fusca var. vacuolata, Chlorella glucotropha, Chlorella infusionum, Chlorella infusionum var. Actophila, Chlorella infusionum var. Auxenophila, Chlorella kessleri, Chlorella luteoviridis, Chlorella luteoviridis var. aureoviridis, Chlorella luteoviridis var. Lutescens, Chlorella miniata, Chlorella minutissima, Chlorella mutabilis, Chlorella nocturna, Chlorella parva, Chlorella photophila, Chlorella pringsheimii, Chlorella protothecoides, Chlorella pyrenoidosa, Chlorella regularis, Chlorella regularis var. minima, Chlorella regularis var. umbricata, Chlorella reisiglii, Chlorella saccharophila, Chlorella saccharophila var. ellipsoidea, Chlorella salina, Chlorella simplex, Chlorella sorokiniana, Chlorella sp., Chlorella sphaerica, Chlorella stigmatophora, Chlorella vanniellii, Chlorella vulgaris, Chlorella vulgaris, Chlorella vulgarisf tertia, Chlorella vulgaris var. airidis, Chlorella vulgaris var. vulgaris, Chlorella vulgaris var. vulgarisf tertia, Chlorella vulgaris var. vulgarisf viridis, Chlorella xanthella, and Chlorella zofingiensis.

67. The method of claim 58, wherein the microalgae are selected from the group consisting of Bracteococcus minor, Chlorella ellipsoidea, Chlorella kessleri, Chlorella luteoviridis, Bracteococcus medionucleatus, Chlorella minutissima, Chlorella ovalis, Chlorella protohecoides, Chlorella saccharophila, Chlorella sorokiniana, Chlorella sp., Chlorella vulgaris, Parachlorella kessleri, Prototheca moriformis, and Pseudochlorella aquatica.

68. A method of manufacturing lipid comprising:

a. providing a microorganism capable of producing lipid;

b. culturing the microorganism in growth media, wherein the growth media comprises depolymerized cellulosic material;

c. allowing the microorganism to accumulate at least 10% of its dry cell weight as lipid; and

d. extracting the lipid from the microorganism.

69. The method of claim 68, wherein the microorganism is an oleaginous yeast.

70. The method of claim 69, wherein the oleaginous yeast is selected from the group consisting of Cryptococcus curvatus, Cryptococcus terricolus, Candida sp., Lipomyces starkeyi, Lipomyces lipofer, Endomycopsis vernalis, Rhodotorula glutinis, Rhodotorula gracilis, and Yarrowia lipolytica.

71. The method of claim 68, wherein the microorganism is a fungus.

72. The method of claim 71, wherein the fungus is selected from the group consisting of a species of the genus Mortierella, Mortierrla vinacea, Mortierella alpine, Pythium debaryanum, Mucor circinelloides, Aspergillus ochraceus, Aspergillus terreus, Pennicillium iilacinum, a species of the genus Hensenulo, a species of the genus Chaetomium, a species of the genus Cladosporium, a species of the genus Malbranchea, a species of the genus Rhizopus, and a species of the genus Pythium.

73. The method of claim 68, wherein the microorganism is a microalgae.

74. The method of claim 73, wherein the microalgae is selected from the group consisting of the microalgae listed in Table 1.

75. The method of claim 73, wherein the microalgae is selected from the group consisting of Chlorella anitrata, Chlorelia antarctica, Chlorella aureoviridis, Chlorella candida, Chlorella capsulata, Chlorella desiccata, Chlorella ellipsoidea, Chlorella emersonii, Chlorellafusca, Chlorellafusca var. vacuolata, Chlorella glucotropha, Chlorella infusionum, Chlorella infusionum var. Actophila, Chlorella infusionum var. Auxenophila, Chlorella kessleri, Chlorella luteoviridis, Chlorella luteoviridis var. aureoviridis, Chlorella luteoviridis var. Lutescens, Chlorella miniata, Chlorella minutissima, Chlorella mutabilis, Chlorella nocturna, Chlorella parva, Chlorella photophila, Chlorella pringsheimii, Chlorella protothecoides, Chlorella pyrenoidosa, Chlorella regularis, Chlorella regularis var. minima, Chlorella regularis var. umbricata, Chlorella reisiglii, Chlorella saccharophila, Chlorella saccharophila var. ellipsoidea, Chlorella salina, Chlorella simplex, Chlorella sorokiniana, Chlorella sp., Chlorella sphaerica, Chlorella stigmatophora, Chlorella vanniellii, Chlorella vulgaris, Chlorella vulgaris, Chlorella vulgaris f. tertia, Chlorella vulgaris var. airidis, Chlorella vulgaris var. vulgaris, Chlorella vulgaris var. vulgarisf tertia, Chlorella vulgaris var. vulgarisf viridis, Chlorella xanthella, and Chlorella zofingiensis.

76. The method of claim 73, wherein the microalgae is selected from the group consisting of Bracteococcus minor, Chlorella ellipsoidea, Chlorella kessleri, Chlorella luteoviridis, Bracteococcus medionucleatus, Chlorella minutissima, Chlorella ovalis, Chlorella protohecoides, Chlorella saccharophila, Chlorella sorokiniana, Chlorella sp., Chlorella vulgaris, Parachlorella kessleri, Prototheca moriformis, and Pseudochlorella aquatica.

77. The method of claim 68, wherein the depolymerized cellulosic material is selected from the group consisting of sugarcane bagasse, rice hulls, corn stover, wheat straw, rice straw, sugar beet pulp, citrus pulp, citrus peels, hardwood thinnings, softwood thinnings, wood chips, sawdust, pulp mill waste, urban paper waste, grass clippings, switchgrass, hybrid poplar wood, miscanthus, fiber cane, and fiber sorghum.

78. The method of claim 68, wherein the extracting the lipid from the microorganism comprises lysing the microorganism and recovering the lipid using an organic solvent.

79. The method of claim 78, wherein the organic solvent is hexane.

80. The method of claim 68, wherein the microorganism is allowed to accumulate at least 25% of its dry cell weight as lipid.

81. The method of claim 68, wherein the microorganism is allowed to accumulate at least 50% of its dry cell weight as lipid.