Granted Patent B2

US 11,421,235 · App. 16/604,425 · Granted Aug 23, 2022

Inventors: Jeffrey David Orth (Alameda, CA); Louis A. Clark (San Francisco, CA); Lily Yuin Chao (San Francisco, CA); Na My Trinh (Walnut Creek, CA); Christopher Cheyney Farwell (South San Francisco, CA); Xinhua Zhao (Dublin, CA); Matthias Helmut Schmalisch (South San Francisco, CA); Grayson Thomas Wawrzyn (Oakland, CA); Xuezhi Li (Hayward, CA); Mark Anton Held (Emeryville, CA); Kevin Lee Dietzel (Pacifica, CA); James Kealey (Sebastopol, CA)

Assignee: PRECIGEN, INC.

C12N15/52C12N9/0006C12N9/1022C12N9/1205C12N9/88C12N15/70C12P7/16C12P7/24C12Y101/01001C12Y101/01086C12Y202/01006C12Y401/01072C12Y402/01009

Abstract

Claims (25)

1. A genetically modified microorganism capable of converting a C 1 carbon to a multicarbon product, wherein the microorganism is Methylococcus capsulatus and comprises:

(a) a heterologous gene encoding an acetolactate synthase (AlsS) comprising the amino acid sequence of SEQ ID NO: 99; and

(b) a heterologous gene encoding a ketol-acid reductoisomerase (KARI) comprising the amino acid sequence of SEQ ID NO: 4;

(c) a heterologous gene encoding a dihydroxy-acid dehydratase (DHAD) comprising the amino acid sequence of SEQ ID NO: 6;

(d) a heterologous gene encoding a 2-keto acid decarboxylase (KDC) comprising the amino acid sequence of SEQ ID NO: 10; and

(e) a heterologous gene encoding an ADH comprising the amino acid sequence of SEQ ID NO: 14.

2. The genetically modified microorganism of claim 1 , further comprising a sugar permease gene.

3. The genetically modified microorganism of claim 1 , wherein at least one of the gene(s) encoding the AlsS, the KARI, the DHAD, the KDC, or the ADH is overexpressed.

4. The genetically modified microorganism of claim 1 , comprising multiple copies of at least one of the gene(s) encoding the AlsS, the KARI, the DHAD, the KDC, and/or the ADH.

5. The genetically modified microorganism of claim 1 , wherein at least one of the gene(s) encoding the AlsS, the KARI, the DHAD, the KDC, or the ADH is under the control of a rare earth metal switch.

6. The genetically modified microorganism of claim 5 , wherein the rare earth metal switch is a lanthanum switch.

7. A method of making the genetically modified microorganism of claim 1 comprising contacting a microorganism with a polynucleotide encoding the AlsS and at least one polynucleotide encoding the KARI, the DHAD, the KDC, and/or the ADH.

8. A method of making the genetically modified microorganism of claim 2 , comprising contacting a microorganism with a polynucleotide encoding a sugar permease.

9. A method of making an alcohol or an aldehyde from a C 1 carbon comprising:

(a) contacting a C 1 carbon with the genetically modified microorganism of claim 1 ; and

(b) growing the genetically modified microorganism to produce the aldehyde or alcohol.

10. A method of making a useful product comprising:

contacting the genetically modified microorganism of claim 1 with a C 1 carbon substrate; and

(b) growing the genetically modified microorganism to produce the useful product, wherein the useful product comprises 2-acetolactate, 2,3-butanediol (2,3-BDO), diacetyl, 2,3-dihydroxy-2-methylbutanoic acid, 2,3-dihydroxyisovalerate, amino acids, ketoisovalerate, isobutyraldehyde, isobutyrate, methyl methacrylate (MMA), isovaleraldehyde, isovalerate, isopentanol, isoamyl acetate, pentadecanoic acid, isobutene, and/or p-xylene.

11. The genetically modified microorganism of claim 1 , wherein:

(a) the heterologous gene encoding the AlsS comprises the nucleic acid sequence of SEQ ID NO: 100;

(b) the heterologous gene encoding the KARI comprises the nucleic acid sequence of SEQ ID NO: 3;

(c) the heterologous gene encoding the DHAD comprises the nucleic acid sequence of SEQ ID NO: 5;

(d) the heterologous gene encoding the KDC comprises the nucleic acid sequence of SEQ ID NO: 9; and

(e) the heterologous gene encoding the ADH comprises the nucleic acid sequence of SEQ ID NO: 13.

Assignments (8)

Continuity (3)