Fatty Acid Desaturases From Primula

• US 20080063691A1
• Filed: 08/20/2004
• Published: 03/13/2008
• Est. Priority Date: 08/21/2003
• Status: Active Grant

• Current Assignee: Monsanto Technology LLC (Bayer AG)
• Original Assignee: Monsanto Technology LLC (Bayer AG)
• Inventors: Ursin, Virginia, Gonzales, Jennifer, Larosa, Thomas J., Dong, Fenggao, Froman, Byron, Screen, Steven E.
• Granted Patent: US 8,173,870 B2
• Time in Patent Office: Days
• US Class Current: 424/442
• CPC Class Codes: A61P 3/02 Nutrients, e.g. vitamins, minerals
• CPC Class Codes: A61P 37/00 Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C12N 15/8247 involving modified lipid metabolism, e.g. seed oil composition
• CPC Class Codes: C12N 9/0004 Oxidoreductases (1.)
• CPC Class Codes: C12P 7/6427 Polyunsaturated fatty acids [PUFA], i.e. having two or more double bonds in their backbone

Abstract

The invention relates generally to methods and compositions concerning desaturase enzymes that modulate the number and location of double bonds in long chain poly-unsaturated fatty acids (LC-PUFA'"'"'s). In particular, the invention relates to methods and compositions for improving omega-3 fatty acid profiles in plant products and parts using desaturase enzymes and nucleic acids encoding for such enzymes. In particular embodiments, the desaturase enzymes are Primula Δ6-desaturases. Also provided are improved soybean oil compositions having SDA and a beneficial overall content of omega-3 fatty acids relative to omega-6 fatty acids.

Claims

1. An isolated polynucleotide encoding a polypeptide having desaturase activity that desaturates a fatty acid molecule at carbon 6, wherein the polynucleotide is selected from the group consisting of:

(a) a polynucleotide encoding the polypeptide sequence of SEQ ID NO;

4, SEQ ID NO;

5, SEQ ID NO;

22, SEQ ID NO;

24, SEQ ID NO;

26, SEQ ID NO;

46 or SEQ ID NO;

48;

(b) a polynucleotide comprising the nucleic acid sequence of SEQ ID NO;

2, SEQ ID NO;

3, SEQ ID NO;

21, SEQ ID NO;

23, SEQ ID NO;

25, SEQ ID NO;

45 or SEQ ID NO;

47;

(c) a polynucleotide hybridizing to SEQ ID NO;

2, SEQ ID NO;

3, SEQ ID NO;

21, SEQ ID NO;

23, SEQ ID NO;

25, SEQ ID NO;

45 or SEQ ID NO;

47, or a complement thereof, under conditions of 5×

SSC, 50% formamide and 42°

C.; and

(d) a polynucleotide encoding a polypeptide with at least 90% sequence identity to a polypeptide sequence of SEQ ID NO;

4, SEQ ID NO;

5, SEQ ID NO;

22, SEQ ID NO;

24, SEQ ID NO;

26, SEQ ID NO;

46 or SEQ ID NO;

48.

3. The isolated polynucleotide of claim 1, further defined as encoding a polypeptide with at least 95% sequence identity to a polypeptide sequence of SEQ ID NO:

4, SEQ ID NO;

5, SEQ ID NO;

22, SEQ ID NO;

24, SEQ ID NO;

26, SEQ ID NO;

46 or SEQ ID NO;

48.

4. The isolated polynucleotide of claim 1, further defined as operably linked to a heterologous promoter.

6. A recombinant vector comprising the isolated polynucleotide sequence of claim 1.

7. The recombinant vector of claim 6, further comprising at least one additional sequence chosen from the group consisting of:

(a) regulatory sequences operatively linked to the polynucleotide;

(b) selection markers operatively linked to the polynucleotide;

(c) marker sequences operatively linked to the polynucleotide;

(d) a purification moiety operatively linked to the polynucleotide; and

(e) a targeting sequence operatively linked to the polynucleotide.

8. The recombinant vector of claim 6, further defined as comprising a promoter operably linked to said isolated polynucleotide.

9. The recombinant vector of claim 8, wherein the promoter is a developmentally-regulated, organelle-specific, tissue-specific, constitutive or cell-specific promoter.

10. The recombinant vector of claim 8, wherein said promoter is selected from the group consisting of 35 S CaMV, 34 S FMV, Napin, 7 S alpha, 7 S alpha′

, Glob, and Lec.

11. The recombinant vector of claim 6, defined as an isolated expression cassette.

12. A transgenic plant transformed with the recombinant vector of claim 6.

13. The transgenic plant of claim 12, further defined as transformed with a nucleic acid sequence encoding a polypeptide having desaturase activity that desaturates a fatty acid molecule at carbon 12.

14. The transgenic plant of claim 12, further defined as transformed with a nucleic acid sequence encoding a polypeptide having desaturase activity that desaturates a fatty acid molecule at carbon 15.

15. A host cell transformed with the recombinant vector of claim 6.

16. The host cell of claim 15, wherein said host cell expresses a protein encoded by said vector.

17. The host cell of claim 15, wherein the cell has inherited said recombinant vector from a progenitor of the cell.

18. The host cell of claim 15, wherein the cell has been transformed with said recombinant vector.

19. The host cell of claim 15, defined as a plant cell.

20. A seed of the plant of claim 12, wherein the seed comprises the recombinant vector.

21. A method of producing seed oil containing omega-3 fatty acids from plant seeds, comprising the steps of:

(a) obtaining seeds of a plant according to claim 12; and

(b) extracting the oil from said seeds.

22. A method of producing a plant comprising seed oil containing altered levels of omega-3 fatty acids comprising introducing the recombinant vector of claim 6 into an oil-producing plant.

23. The method of claim 22, wherein introducing the recombinant vector comprises plant breeding.

24. The method of claim 22, wherein introducing the recombinant vector comprises genetic transformation.

25. The method of claim 22, wherein the plant is a plant selected from the group consisting of Arabidopsis thaliana, oilseed Brassica, rapeseed, sunflower, safflower, canola, corn, soybean, cotton, flax, jojoba, Chinese tallow tree, tobacco, cocoa, peanut, fruit plants, citrus plants, and plants producing nuts and berries.

26. The method of claim 22, wherein the plant is further defined as transformed with a nucleic acid sequence encoding a polypeptide having desaturase activity that desaturates a fatty acid molecule at carbon 15.

27. The method of claim 26, wherein stearidonic acid is increased.

28. The method of claim 22, further defined as comprising introducing the recombinant vector of claim 6 into a plurality of oil-producing plants and screening said plants or progeny thereof having inherited the recombinant vector for a plant having a desired profile of omega-3 fatty acids.

2. (canceled)

5. An isolated polypeptide comprising the polypeptide sequence of SEQ ID NO:

4, SEQ ID NO;

5, SEQ ID NO;

22, SEQ ID NO;

24, SEQ ID NO;

26, SEQ ID NO;

46 or SEQ ID NO;

48, or a fragment thereof having desaturase activity that desaturates a fatty acid molecule at carbon 6.

29. An endogenous soybean seed oil having a stearidonic acid content of from about 5% to about 50% and a gamma-linoleic acid content of less than 10%.

30. The soybean seed oil of claim 29, further defined as comprising less than 10% combined alpha-linoleic acid, linoleic acid and gamma-linoleic acid.

31. The soybean seed oil of claim 29, wherein the stearidonic acid content is further defined as selected from the group consisting of from about 15% to about 35% and from about 22% to about 30%.

33. The soybean seed oil of claim 29, wherein the gamma-linoleic acid is further defined as less than 5%.

34. The soybean seed oil of claim 29, wherein the stearidonic acid content is further defined as from about 15% to about 35% and the gamma-linoleic acid content is further defined as less than 5%.

35. The soybean seed oil of claim 29, wherein the ratio of omega-3 to omega-6 fatty acids in the oil is selected from the group consisting of from about 0.35:

1 to about 3.5;

1 and from about 1;

1 to about 3.5;

1.

37. A method of increasing the nutritional value of an edible product for human or non-human animal consumption, comprising adding the soybean seed oil of claim 29 to the edible product.

38. The method of claim 37, wherein the edible product is selected from the group consisting of human food, animal feed and a food supplement.

39. The method of claim 37, wherein the soybean seed oil increases the stearidonic acid content of the edible product.

40. The method of claim 37, wherein the soybean seed oil increases the ratio of omega-3 to omega-6 fatty acids of the edible product.

41. The method of claim 37, wherein the edible product lacks stearidonic acid prior to adding the soybean seed oil.

42. A method of manufacturing food and/or feed, comprising adding the soybean seed oil of claim 29 to starting food and/or feed ingredients to produce the food and/or feed.

43. Food or feed made by the method of claim 42.

44. A method of providing stearidonic acid to a human or non-human animal, comprising administering the soybean seed oil of claim 29 to said human or non-human animal.

45. The method of claim 44, wherein the soybean seed oil is administered in an edible composition.

46. The method of claim 45, wherein the edible composition is food or feed.

47. The method of claim 46, wherein the food comprises beverages, infused foods, sauces, condiments, salad dressings, fruit juices, syrups, desserts, icings and fillings, soft frozen products, confections or intermediate food.

48. The method of claims 45, wherein the edible composition is substantially a liquid or a solid, a food supplement or a nutraceutical.

50. The method of claim 44, wherein the soybean seed oil is administered to a human, a non-human animal, livestock or poultry.

32. (canceled)

36. (canceled)

49. (canceled)

51-52. -52. (canceled)