Tailored oils produced from recombinant heterotrophic microorganisms

• US 9,255,282 B2
• Filed: 05/13/2014
• Issued: 02/09/2016
• Est. Priority Date: 05/28/2010
• Status: Active Grant

• Current Assignee: Corbion Biotech, Inc. (Corbion NV)
• Original Assignee: Solazyme Inc.
• Inventors: Franklin, Scott, Somanchi, Aravind, Wee, Janice, Rudenko, George, Moseley, Jeffrey L., Rakitsky, Walt
• Primary Examiner(s): Monshipouri, Maryam
• Application Number:
  US14/276,943
• Publication Number:
  US 20140315267A1
• Time in Patent Office: 637 Days
• US Class Current: 1/1
• CPC Class Codes: A23D 7/00 Edible oil or fat compositions containing an aqueous phase, e.g. margarines
• CPC Class Codes: A23D 9/00 Other edible oils or fats, e.g. shortenings, cooking oils
• CPC Class Codes: A23K 20/158 Fatty acids; Fats; Products containing oils or fats
• CPC Class Codes: C11B 1/00 Production of fats or fatty oils from raw materials
• 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 15/67 General methods for enhancing the expression
• CPC Class Codes: C12N 15/79 Vectors or expression systems specially adapted for eukaryotic hosts
• CPC Class Codes: C12N 15/8216 Methods for controlling, regulating or enhancing expression of transgenes in plant cells
• CPC Class Codes: C12N 9/0071 acting on paired donors with incorporation of molecular oxygen (1.14)
• CPC Class Codes: C12N 9/16 acting on ester bonds (3.1)
• CPC Class Codes: C12N 9/2431 Beta-fructofuranosidase (3.2.1.26), i.e. invertase
• CPC Class Codes: C12N 9/2465 acting on alpha-galactose-glycoside bonds, e.g. alpha-galactosidase (3.2.1.22)
• CPC Class Codes: 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
• CPC Class Codes: C12P 7/6445 Glycerides
• 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: C12Y 114/19001 Stearoyl-CoA 9-desaturase (1.14.19.1), i.e. DELTA9-desaturase
• CPC Class Codes: C12Y 301/02 Thioester hydrolases (3.1.2)
• CPC Class Codes: C12Y 301/02014 Oleoyl-[acyl-carrier-protein] hydrolase (3.1.2.14), i.e. ACP-thioesterase
• CPC Class Codes: C12Y 302/01022 Alpha-galactosidase (3.2.1.22)
• 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: Y02P 30/20 : using bio-feedstock
• CPC Class Codes: Y02T 50/678 : Aviation using fuels of non-fossil origin

Abstract

Methods and compositions for the production of oil, fuels, oleochemicals, and other compounds in recombinant microorganisms are provided, including oil-bearing microorganisms and methods of low cost cultivation of such microorganisms. Microalgal cells containing exogenous genes encoding, for example, a lipase, a sucrose transporter, a sucrose invertase, a fructokinase, a polysaccharide-degrading enzyme, a keto acyl-ACP synthase enzyme, a fatty acyl-ACP thioesterase, a fatty acyl-CoA/aldehyde reductase, a fatty acyl-CoA reductase, a fatty aldehyde reductase, a fatty aldehyde decarbonylase, and/or an acyl carrier protein are useful in manufacturing transportation fuels such as renewable diesel, biodiesel, and renewable jet fuel, as well as oleochemicals such as functional fluids, surfactants, soaps and lubricants.

Claims

1. A recombinant eukaryotic host cell comprising an exogenous nucleic acid encoding a THIC enzyme, wherein the recombinant eukaryotic host cell expresses the nucleic acid and is capable of growing in a thiamine-free culture medium.

2. The recombinant eukaryotic host cell of claim 1, wherein the exogenous nucleic acid encodes a Coccomyxa THIC enzyme, an Arabidopsis THIC enzyme, or a Synechocystis THIC enzyme.

3. The recombinant eukaryotic host cell of claim 1 further comprising a second nucleic acid encoding an enzyme selected from the group consisting of sucrose invertase, thioesterase, ketoacyl synthase and desaturase.

4. The recombinant eukaryotic host cell of claim 3, wherein the second nucleic acid encodes a sucrose invertase.

5. The recombinant eukaryotic host cell of claim 1, wherein the eukaryotic host cell further comprises nucleic acids that encode a sucrose invertase and a thioesterase.

6. The recombinant eukaryotic host cell of claim 1, wherein the eukaryotic host cell further comprises nucleic acids that encode a sucrose invertase and a desaturase.

7. The recombinant eukaryotic host cell of claim 1, wherein the eukaryotic host cell further comprises nucleic acids that encode a sucrose invertase, a thioesterase, a desaturase and optionally a ketoacyl synthase.

8. The recombinant eukaryotic host cell of claim 1, wherein the recombinant eukaryotic host cell comprises at least 40% lipid by dry cell weight.

9. The recombinant eukaryotic host cell of claim 8, wherein the eukaryotic host cell is an oleaginous microorganism.

10. The recombinant eukaryotic host cell of claim 9, wherein the host cell is a microalga.

11. The recombinant eukaryotic host cell of claim 10, wherein the microalga is Prototheca or Chlorella.

12. The recombinant eukaryotic host cell of claim 11, wherein the microalga is Prototheca moriformis.

13. A method of cultivating a recombinant eukaryotic host cell, the host cell comprising an exogenous nucleic acid encoding a THIC enzyme, wherein the recombinant eukaryotic host cell expresses the nucleic acid and is capable of growing in a thiamine-free culture medium, the method comprising the steps of:

a. inoculating the recombinant eukaryotic host cell into a culture medium, wherein the culture medium is essentially free of thiamine;

b. cultivating the host cell to a desired cell density; and

c. harvesting the host cell from the culture medium.

14. The method of claim 13, wherein the eukaryotic host cell is an oleaginous microorganism.

15. The method of claim 14, wherein the host cell is a microalga.

16. The method of claim 15, wherein the microalga is Prototheca or Chlorella.

17. The method of claim 16, wherein the microalga is Prototheca moriformis.