# US 7,745,694 B1 · Methods and compositions for synthesis of long chain polyunsaturated fatty acids in plants > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 7,745,694 B1 - Document ID: US7745694B1 - Status: Granted - Kind: B1 - Application number: 09/355,903 - Filed: 1998-04-10 - Granted: 2010-06-29 - Earliest claimed: 1997-04-11 - Assignees: Monsanto Technology LLC - Inventors: Deborah Knutzon; Pradip Mukerji; Yung-Sheng Huang; Jennifer Thurmond; Sunita Chaudhary; Amanda Eun-Yeong Leonard - Web page: https://patentgrep.com/patent/US7745694B1 ## CPC - C12N 9/0083: {Miscellaneous (1.14.99)} - A23D 9/00: Other edible oils or fats, e.g. shortenings or cooking oils (preservation of other edible oils A23B20/30) - A23K 20/158: Fatty acids; Fats; Products containing oils or fats - A23L 33/12: Fatty acids or derivatives thereof - A23L 33/21: Addition of substantially indigestible substances, e.g. dietary fibres - A23L 33/40: {Complete food formulations for specific consumer groups or specific purposes, e.g. infant formula} - A61P 3/02: Nutrients, e.g. vitamins, minerals - C12N 15/81: for yeasts - C12N 15/8247: {involving modified lipid metabolism, e.g. seed oil composition} - C12P 7/64: Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats - C12P 7/6427: Polyunsaturated fatty acids [PUFA], i.e. having two or more double bonds in their backbone - C12P 7/6431: Linoleic acids [18:2[n-6]] - C12P 7/6472: containing polyunsaturated fatty acid [PUFA] residues, i.e. having two or more double bonds in their backbone - A01K 2217/05: Animals comprising random inserted nucleic acids (transgenic) - A23V 2002/00: Food compositions, function of food ingredients or processes for food or foodstuffs - A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00) - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) - A23D: EDIBLE OILS OR FATS, e.g. MARGARINES, SHORTENINGS OR COOKING OILS (preservation thereof A23B20/00; production, refinement or preservation of animal or vegetable fats or oils C11B, C11C) - A23K: FODDER - A23L: FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF (preservation thereof A23B) - A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS - C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)} - A01K: ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS - A23V: INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION - A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D) ## Abstract The present invention relates to compositions and methods for preparing polyunsaturated long chain fatty acids in plants, plant parts and plant cells, such as leaves, roots, fruits and seeds. Nucleic acid sequences and constructs encoding fatty acid desaturases, including Δ5-desaturases, Δ6-desaturases and Δ12-desaturases, are used to generate transgenic plants, plant parts and cells which contain and express one or more transgenes encoding one or more desaturases. Expression of the desaturases with different substrate specificities in the plant system permit the large scale production of polyunsaturated long chain fatty acids such as docosahexaenoic acid, eicosapentaenoic acid, α-linolenic acid, gamma-linolenic acid, arachidonic acid and the like for modification of the fatty acid profile of plants, plant parts and tissues. Manipulation of the fatty acid profiles allows for the production of commercial quantities of novel plant oils and products. ## Claims ### Claim 1 A method for producing a plant comprising a tissue and/or part thereof with an altered fatty acid profile comprising: growing a plant comprising a tissue and/or part thereof comprising a plurality of cells, each of said plurality comprising a nucleic acid comprising the sequence depicted in SEQ ID NO:1, said nucleic acid operably linked to transcription and translation control signals functional in said cells, wherein a polypeptide encoded by said nucleic acid is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile thereof. ### Claim 2 (depends on 1) The method of claim 1, wherein said tissue and/or part is a seed of said plant. ### Claim 3 (depends on 1) The method of claim 1, wherein said tissue and/or part is a leaf of said plant. ### Claim 4 (depends on 1) The method of claim 1, wherein said tissue and/or part is a fruit of said plant. ### Claim 5 (depends on 1) The method of claim 1, wherein said tissue and/or part is a flower of said plant. ### Claim 6 (depends on 1) The method of claim 1, wherein said tissue and/or part is a root of said plant. ### Claim 7 (depends on 1) The method of claim 1, wherein at least one of said transcription and translation control signals is endogenous to said plant. ### Claim 8 A method for producing a plant comprising a tissue and/or part thereof with an altered fatty acid profile comprising: growing a plant comprising a tissue and/or part thereof comprising a plurality of cells, each of said plurality comprising a nucleic acid with at least 95% homology to the sequence depicted in SEQ ID NO:1, said nucleic acid operably linked to transcription and translation control signals function in said cells, wherein a polypeptide encoded by said nucleic acid forms a monounsaturated bond between carbons 6 and 7 of a fatty acid as numbered from a carboxy terminus thereof, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile thereof. ### Claim 9 (depends on 8) The method of claim 8, wherein said tissue and/or part is a seed of said plant. ### Claim 10 (depends on 8) The method of claim 8, wherein said tissue and/or part is a leaf of said plant. ### Claim 11 (depends on 8) The method of claim 8, wherein said tissue and/or part is a fruit of said plant. ### Claim 12 (depends on 8) The method of claim 8, wherein said tissue and/or part is a flower of said plant. ### Claim 13 (depends on 8) The method of claim 8, wherein said tissue and/or part is a root of said plant. ### Claim 14 (depends on 8) The method of claim 8, wherein at least one of said transcription and translation control signals is endogenous to said plant. ### Claim 15 A method for producing a plant comprising a tissue and/or part thereof with an altered fatty acid profile comprising: growing a plant comprising a tissue and/or plant thereof comprising a plurality of cells, each of said plurality comprising a nucleic acid operably linked to transcription and translation control signals functional in said cells, wherein said nucleic acid encodes a deletion mutant of the polypeptide depicted in SEQ ID NO:2, wherein a polypeptide encoded by said nucleic acid forms a monounsaturated bond between carbons 6 and 7 of a fatty acid as numbered from a carboxy terminus thereof, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile thereof. ### Claim 16 A method for producing a plant comprising a tissue and/or part thereof with an altered fatty acid profile comprising: growing a plant comprising a tissue and/or part thereof comprising a plurality of cells, each of said plurality comprising a polypeptide comprising the amino acid sequence depicted in SEQ ID NO:2, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile thereof. ### Claim 17 (depends on 16) The method of claim 16, wherein said tissue and/or part is a seed of said plant. ### Claim 18 (depends on 16) The method of claim 16, wherein said tissue and/or part is a leaf of said plant. ### Claim 19 (depends on 16) The method of claim 16, wherein said tissue and/or part is a fruit of said plant. ### Claim 20 (depends on 16) The method of claim 16, wherein said tissue and/or part is a flower of said plant. ### Claim 21 (depends on 16) The method of claim 16, wherein said tissue and/or part is a root of said plant. ### Claim 22 A method for producing a plant comprising a tissue and/or part thereof with an altered fatty acid profile comprising: growing a plant comprising a tissue and/or part thereof comprising a plurality of cells, each of said plurality comprising a polypeptide with at least 95% homology to the sequence depicted in SEQ ID NO:2, wherein said polypeptide forms a monounsaturated bond between carbons 6 and 7 of a fatty acid as numbered from a carboxy terminus thereof, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile thereof. ### Claim 23 A method for producing a plant comprising a tissue and/or part thereof with an altered fatty acid profile comprising: growing a plant comprising a tissue and/or part thereof comprising a plurality of cells, each of said plurality comprising a nucleic acid that hybridizes preferentially to a complement of the sequence depicted in SEQ ID NO:1 under hybridization conditions suitable for selectively screening a recombinant DNA library using a probe comprising said complement, said recombinant DNA library comprising sequences obtained from a Mortierella species, said nucleic acid operably linked to transcription and translation control signals functional in said cells, wherein a polypeptide encoded by said nucleic acid forms a monounsaturated bond between carbons 6 and 7 of a fatty acid as numbered from a carboxy terminus thereof, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile thereof. ### Claim 24 A method for producing oil with an altered fatty acid profile comprising extracting said oil with an altered fatty acid profile from at least a plant tissue and/or part produced by growing a plant comprising the tissue and/or part, said tissue and/or part comprising a plurality of cells, each of said plurality comprising a nucleic acid comprising the sequence depicted in SEQ ID NO:1, said nucleic acid operably linked to transcription and translation control signals functional in said cells, wherein a polypeptide encoded by said nucleic acid is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile of said oil, wherein the fatty acid profile is altered as compared to that of oil extracted from a parental strain of said plant not expressing said polypeptide. ### Claim 25 (depends on 24) The method of claim 24, further comprising purifying a component of said oil. ### Claim 26 (depends on 25) The method of claim 25, wherein said component is a phospholipid. ### Claim 27 (depends on 25) The method of claim 25, wherein said component is a sulfolipid. ### Claim 28 (depends on 25) The method of claim 25, wherein said component is a glycolipid. ### Claim 29 (depends on 25) The method of claim 25, wherein said component is a acylglycerol. ### Claim 30 (depends on 29) The method of claim 29, wherein said component is a monoacylglycerol. ### Claim 31 (depends on 29) The method of claim 29, wherein said component is a diacylglycerol. ### Claim 32 (depends on 29) The method of claim 29, wherein said component is a triacylglycerol. ### Claim 33 A method for producing oil with an altered fatty acid profile comprising extracting said oil from at least a plant tissue and/or part produced by growing a plant comprising the tissue and/or part, said tissue and/or part comprising a plurality of cells, each of said plurality comprising a nucleic acid with at least 95% homology to the sequence depicted in SEQ ID NO:1, said nucleic acid operably linked to transcription and translation control signals function in said cells, wherein a polypeptide encoded by said nucleic acid forms a monounsaturated bond between carbons 6 and 7 of a fatty acid as numbered from a carboxy terminus thereof, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile of said oil, wherein the fatty acid profile is altered as compared to that of oil extracted from a parental strain of said plant not expressing said polypeptide. ### Claim 34 A method for producing oil with an altered fatty acid profile comprising extracting said oil with an altered fatty acid profile from at least a plant tissue and/or part produced by growing a plant comprising the tissue and/or part, said tissue and/or part comprising a plurality of cells, each of said plurality comprising a polypeptide comprising the amino acid sequence depicted in SEQ ID NO:2, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile of said oil, wherein the fatty acid profile is altered as compared to that of oil extracted from a parental strain of said plant not expressing said polypeptide. ### Claim 35 A method for producing oil with an altered fatty acid profile comprising extracting said oil with an altered fatty acid profile from at least a plant tissue and/or part produced by growing a plant comprising the tissue and/or part, said tissue and/or part comprising a plurality of cells, each of said plurality comprising a polypeptide with at least 95% homology to the sequence depicted in SEQ ID NO:2, said polypeptide forms a monounsaturated bond between carbons 6 and 7 of a fatty acid as numbered from a carboxy terminus thereof, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile of said oil, wherein the fatty acid profile is altered as compared to that of oil extracted from a parental strain of said plant not expressing said polypeptide. ### Claim 36 A plant comprising a tissue and/or part thereof comprising a plurality of cells, each of said plurality comprising a nucleic acid comprising the sequence depicted in SEQ ID NO:1, said nucleic acid operably linked to transcription and translation control signals functional in said cells, wherein a polypeptide encoded by said nucleic acid is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile of said oil, wherein the fatty acid profile is altered as compared to that of oil extracted from a parental strain of said plant not expressing said polypeptide. ### Claim 37 (depends on 36) A seed isolated from the plant of claim 36. ### Claim 38 (depends on 36) A leaf isolated from the plant of claim 36. ### Claim 39 (depends on 36) A fruit isolated from the plant of claim 36. ### Claim 40 (depends on 36) A flower isolated from the plant of claim 36. ### Claim 41 (depends on 36) A root isolated from the plant of claim 36. ### Claim 42 A plant comprising a tissue and/or part thereof comprising a plurality of cells, each of said plurality comprising a nucleic acid with at least 95% homology to the sequence depicted in SEQ ID NO:1, said nucleic acid operably linked to transcription and translation control signals functional in said tissue and/or part, wherein a polypeptide encoded by said nucleic acid forms a monounsaturated bond between carbons 6 and 7 of a fatty acid as numbered from a carboxy terminus thereof, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile of an oil of said tissue and/or part, wherein the fatty acid profile is altered as compared to that of oil extracted from a parental strain of said plant not expressing said polypeptide. ### Claim 43 (depends on 42) A seed isolated from the plant of claim 42. ### Claim 44 (depends on 42) A leaf isolated from the plant of claim 42. ### Claim 45 (depends on 42) A fruit isolated from the plant of claim 42. ### Claim 46 (depends on 42) A flower isolated from the plant of claim 42. ### Claim 47 (depends on 42) A root isolated from the plant of claim 42. ### Claim 48 A plant comprising a tissue and/or part thereof comprising a plurality of cells, each of said plurality comprising a polypeptide comprising the amino acid sequence depicted in SEQ ID NO:2, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile of said oil, wherein the fatty acid profile is altered as compared to that of oil extracted from a parental strain of said plant not expressing said polypeptide. ### Claim 49 (depends on 48) A seed isolated from the plant of claim 48. ### Claim 50 (depends on 48) A leaf isolated from the plant of claim 48. ### Claim 51 (depends on 48) A fruit isolated from the plant of claim 48. ### Claim 52 (depends on 48) A flower isolated from the plant of claim 48. ### Claim 53 (depends on 48) A root isolated from the plant of claim 48. ### Claim 54 A plant comprising a tissue and/or part thereof comprising a plurality of cells, each of said plurality comprising a polypeptide with at least 95% homology to the sequence depicted in SEQ ID NO:2, wherein said polypeptide forms a monounsaturated bond between carbons 6 and 7 of a fatty acid as numbered from a carboxy terminus thereof, wherein said polypeptide is expressed in sufficient amount in said tissue and/or part to alter the fatty acid profile of an oil of said tissue and/or part, wherein the fatty acid profile is altered as compared to that of oil extracted from a parental strain of said plant not expressing said polypeptide. ### Claim 55 (depends on 54) A seed isolated from the plant of claim 54. ### Claim 56 (depends on 54) A leaf isolated from the plant of claim 54. ### Claim 57 (depends on 54) A fruit isolated from the plant of claim 54. ### Claim 58 (depends on 54) A flower isolated from the plant of claim 54. ### Claim 59 (depends on 54) A root isolated from the plant of claim 54. ### Claim 60 A method for producing oil with an increased amount of a polyunsaturated fatty acid and/or a conjugated form thereof, comprising extracting said oil from at least a plant tissue and/or part produced by growing a plant comprising the tissue and/or part, said tissue and/or part comprising a plurality of cells, each of said plurality comprising a nucleic acid comprising the sequence depicted in SEQ ID NO:1, said nucleic acid operably linked to transcription and translation control signals functional in said cells, wherein a polypeptide encoded by said nucleic acid is expressed in said tissue and/or part and is used to produce said increased amount of the polyunsaturated fatty acid and/or conjugated form of said oil, wherein the fatty acid profile is altered as compared to that of oil extracted from a parental strain of said plant not expressing said polypeptide. ### Claim 61 (depends on 54) A method for producing oil with an increased amount of a polyunsaturated fatty acid and/or a conjugated form thereof, comprising extracting said oil from at least a plant tissue and/or part produced by growing the plant of claim 54.