Method for Producing Unsaturated Omega-3-Fatty Acids in Transgenic Organisms

• US 20070224661A1
• Filed: 02/23/2005
• Published: 09/27/2007
• Est. Priority Date: 02/27/2004
• Status: Active Grant

• Current Assignee: BASF Plant Science GmbH (BASF SE)
• Original Assignee: BASF Plant Science LLC (BASF SE)
• Inventors: Cirpus, Petra, Zank, Thorsten, Heinz, Ernst, Bauer, Jorg
• Granted Patent: US 7,777,098 B2
• Time in Patent Office: Days
• US Class Current: 435/69.100
• 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: C12N 9/0071 acting on paired donors with incorporation of molecular oxygen (1.14)

Abstract

The present invention relates to a process for production of unsaturated ω-3-fatty acids and to a process for production of triglycerides with an elevated content of unsaturated fatty acids, especially of ω-3-fatty acids having more than three double bonds. The invention relates to the production of a transgenic organism, preferably of a transgenic plant or of a transgenic microorganism, with an elevated content of unsaturated ω-3-fatty acids, oils or lipids having ω-3-double bonds as the result of the expression of an ω-3-desaturase from fungi of the family Pythiaceae such as the genus Phytophtora, for example of the genus and species Phytophtora infestans. The invention furthermore relates to the nucleic acid sequences, nucleic acid constructs, vectors and organisms comprising at least one nucleic acid sequence according to the invention, at least one vector comprising the nucleic acid sequence and/or the nucleic acid constructs, and transgenic organisms comprise the abovementioned nucleic acid sequences, nucleic acid constructs and/or vectors. A further part of the invention relates to oils, lipids and/or fatty acids produced by the process according to the invention, and to their use. The invention moreover relates to unsaturated fatty acids and triglycerides with an elevated content of unsaturated fatty acids and their use.

Claims

1. A process for production of compounds of the general formula I

2. The process according to claim 1, wherein in addition to the nucleic acid sequence introduced in item a), which codes for an ω

-3-desaturase activity, further nucleic acid sequences which code for polypeptides with Δ

9-elongase, Δ

6-desaturase, Δ

8-desaturase, Δ

6-elongase, Δ

5-desaturase, Δ

5-elongase or Δ

4-desaturase activity are introduced.

3. The process according to claim 1, wherein the substituents R² or R³ independently of one another are saturated or unsaturated C₁₈-C₂₂-alkylcarbonyl.

4. The process according to claim 1, wherein the substituents R² or R³ independently of one another are unsaturated C₁₈-, C₂₀- or C₂₂-alkylcarbonyl with at least two double bonds.

5. The process according to claim 1, wherein the transgenic organism is a transgenic microorganism or a transgenic plant.

6. The process according to claim 1, wherein the transgenic organism is an oil-producing plant, a vegetable plant or an ornamental.

7. The process according to claim 1, wherein the transgenic organism is a transgenic plant selected from the group of the plant families Adelotheciaceae, Anacardiaceae, Asteraceae, Apiaceae, Betulaceae, Boraginaceae, Brassicaceae, Bromeliaceae, Caricaceae, Cannabaceae, Convolvulaceae, Chenopodiaceae, Crypthecodiniaceae, Cucurbitaceae, Ditrichaceae, Elaeagnaceae, Ericaceae, Euphorbiaceae, Fabaceae, Geraniaceae, Gramineae, Juglandaceae, Lauraceae, Leguminosae, Linaceae or Prasinophyceae.

8. The process according to claim 1, wherein the compounds of the general formula I are isolated from the organism in the form of their oils, lipids or free fatty acids.

9. The process according to claim 1, wherein the compounds of the general formula I are isolated in a concentration of at least 5% by weight based on the total lipid content of the transgenic organism.

10. An oil, lipid or fatty acid or a fraction thereof, produced by the process according to claim 1.

11. An oil, lipid or fatty acid composition which comprises PUFAs produced by a process according to claim 1 and which originates from transgenic plants.

12. A process for the production of oils, lipids or fatty acid compositions comprising mixing oil, lipids or fatty acids according to claim 10 with oils, lipids or fatty acids of animal origin.

13. Feed, foodstuffs, cosmetics or pharmaceuticals prepared from the oil, lipids or fatty acids or a fraction thereof according to claim 10.

14. An isolated nucleic acid sequence which codes for polypeptides with ω

-3-desaturase activity, selected from the group consisting of;

a) a nucleic acid sequence with the sequence shown in SEQ ID NO;

1,

b) nucleic acid sequences which, as the result of the degeneracy of the genetic code, can be derived from the amino acid sequence shown in SEQ ID NO;

2, or

c) derivatives of the nucleic acid sequence shown in SEQ ID NO;

1 which encode polypeptides with at least 60% identity at the amino acid level with SEQ ID NO;

2 and which have ω

-3-desaturase activity.

15. The isolated nucleic acid sequence according to claim 14, wherein the sequence is derived from an alga, a fungus, a microorganism or a nonhuman animal.

16. The isolated nucleic acid sequence according to claim 14, wherein the sequence is derived from the family Pythiaceae.

17. An amino acid sequence which is encoded by the isolated nucleic acid sequence according to claim 14.

18. A gene construct comprising the isolated nucleic acid according to claim 14, wherein the nucleic acid is linked functionally to one or more regulatory signals.

19. The gene construct according to claim 18, wherein the nucleic acid construct comprises additional biosynthesis genes of the fatty acid or lipid metabolism selected from the group acyl-CoA dehydrogenase(s), acyl-ACP [=acyl carrier protein] desaturase(s), acyl-ACP thioesterase(s), fatty acid acyltransferase(s), acyl-CoA:

lysophospholipid acyltransferase(s), fatty acid synthase(s), fatty acid hydroxylase(s), acetyl-coenzyme A carboxylase(s), acyl-coenzyme A oxidase(s), fatty acid desaturase(s), fatty acid acetylenases, lipoxygenases, triacylglycerol lipases, allenoxide synthases, hydroperoxide lyases or fatty acid elongase(s).

20. The gene construct according to claim 18 wherein the nucleic acid construct additionally comprises biosynthesis genes of the fatty acid or lipid metabolism selected from the group Δ

4-desaturase, Δ

5-desaturase, Δ

6-desaturase, Δ

8-desatuase, Δ

9-desaturase, Δ

12-desaturase, Δ

6-elongase, Δ

5-elongase or Δ

9-elongase.

21. A vector comprising the nucleic acid according to claim 14.

22. A transgenic nonhuman organism comprising at least one nucleic acid according to claim 14.

23. The transgenic nonhuman organism according to claim 22, wherein the organism is a microorganism, a nonhuman animal or a plant.

24. The transgenic nonhuman organism according to claim 22, wherein the organism is a plant.