Algal mutants having a locked-in high light acclimated phenotype

Patent metadata

Field	Value
Publication number	US-9982272-B2
Application number	US-201314099879-A
Country	US
Kind code	B2
Filing date	Dec 6, 2013
Priority date	Dec 6, 2012
Publication date	May 29, 2018
Grant date	May 29, 2018

Abstract

Mutant photosynthetic microorganisms having reduced chlorophyll and increased photosynthetic efficiency are provided. The mutants have a locked in high light-acclimated phenotype, in which many of the photosynthetic parameters characteristic of high light acclimated wild type cells are found in the LIHLA mutants when acclimated to low light, such as reduced chlorophyll, reduced NPQ, higher qP, higher E k , higher P max per unit chlorophyll with little to no reduction in P max per cell, and higher rates of electron transport through photosystem II over a wide range of light intensities. Provided herein are constructs for attenuating or disrupting genes are provided for generating mutants having the LIHLA phenotype. Also provided are methods of culturing LIHLA mutants for the production of biomass or other products.

First claim

We claim:

1. An algal mutant deregulated in acclimation to low light, wherein said algal mutant is derived from classical mutagenesis or genetic engineering and comprises:
(a) a disrupted gene encoding a polypeptide having at least 85% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 4 or the amino acid sequence of SEQ ID NO: 8;
(b) a disrupted gene encoding a polypeptide having at least 85% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 6 or the amino acid sequence of SEQ ID NO: 21; or
(c) a disrupted gene encoding a polypeptide having at least 85% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 63 or the amino acid sequence of SEQ ID NO: 66; and further wherein said algal mutant exhibits:
a reduction in chlorophyll under low light conditions, and
higher photochemical quenching (qP) at all physiologically relevant irradiances above 300 μE*m −2 *s −1 ,
with respect to a wild type alga of the same strain.

2. The algal mutant of claim 1 , wherein said reduction in chorophyll is at least a 20% reduction with respect to a wild type alga of the same strain.

3. The algal mutant of claim 2 , wherein said reduction in chlorophyll is at least a 30% reduction with respect to a wild type alga of the same strain.

4. The algal mutant of claim 1 , wherein said algal mutant exhibits onset of nonphotochemical quenching (NPQ) at a higher irradiance than a wild type alga of the same strain.

5. The algal mutant of claim 1 , wherein said algal mutant exhibits lower NPQ at all physiologically relevant irradiances above 400 μE*m −2 *s −1 than a wild type alga of the same strain.

6. The algal mutant of claim 1 , wherein said algal mutant has at least 70% of the maximal photosynthetic rate (P max ) per cell as a wild type alga of the same strain.

7. The algal mutant of claim 1 , wherein said algal mutant has a higher saturating irradiance for photosynthesis (Ek) than a wild type alga of the same strain.

8. The algal mutant of claim 1 , wherein a culture of said algal mutant has greater penetration of light into said culture than does a culture of a wild type alga of the same strain.

9. The algal mutant of claim 1 , wherein said algal mutant has been generated by UV irradiation, gamma irradiation, or chemical mutagenesis.

10. The algal mutant of claim 1 , wherein said algal mutant is a genetically engineered algal mutant.

11. The algal mutant of claim 10 , wherein said algal mutant has been genetically engineered by a means for disrupting a gene encoding a regulator of light acclimation.

12. The algal mutant of claim 11 , wherein said algal mutant has been genetically engineered by insertional mutagenesis.

13. The algal mutant of claim 1 , wherein said algal mutant belongs to a genus selected from the group consisting of: Achnanthes, Amphiprora, Amphora, Ankistrodesmus, Asteromonas, Boekelovia, Bolidomonas, Borodinella, Botrydium, Botryococcus, Bracteococcus, Chaetoceros, Carteria, Chlamydomonas, Chlorococcum, Chlorogonium, Chlorella, Chroomonas, Chrysosphaera, Cricosphaera, Crypthecodinium, Cryptomonas, Cyclotella, Dunaliella, Ellipsoidon, Emiliania, Eremosphaera, Ernodesmius, Euglena, Eustigmatos, Franceia, Fragilaria, Gloeothamnion, Haematococcus, Halocafeteria, Heterosigma, Hymenomonas, Isochrysis, Lepocinclis, Micractinium, Monodus, Monoraphidium, Nannochloris, Nannochloropsis, Navicula, Neochloris, Nephrochloris, Nephroselmis, Nitzschia, Ochromonas, Oedogonium, Oocystis, Ostreococcus, Pavlova, Parachlorella, Pascheria, Pelagomonas, Phaeodactylum, Phagus, Picochlorum, Platymonas, Pleurochrysis, Pleurococcus, Prototheca, Pseudochlorella, Pseudoneochloris, Pseudostaurastrum, Pyramimonas, Pyrobotrys, Scenedesmus, Skeletonema, Spyrogyra, Stichococcus, Tetraselmis, Thalassiosira, Tribonema, Vaucheria, Viridiella, Vischeria , and Volvox.

14. The algal mutant of claim 13 , wherein said algal mutant belongs to a genus selected from the group consisting of: Amphora, Chaetoceros, Cyclotella, Ellipsoidon, Fragilaria, Monodus, Nannochloropsis, Navicula, Nitzschia, Phaeodactylum , and Thalassiosira.

15. An algal biomass comprising the algal mutant of claim 1 .

16. A method of producing an algal product, comprising culturing the algal mutant of claim 1 and isolating at least product produced by said algal mutant from said culture.

17. The method of claim 16 , wherein said product is algal biomass.

18. The method of claim 16 , wherein said product is a lipid, a protein, a peptide, one or more amino acids, a carbohydrate, one or more nucleotides, a vitamin, a cofactor, a hormone, an antioxidant, or a pigment or colorant.

19. The method of claim 16 , wherein said product is a lipid.

20. The method of claim 19 , wherein said algal mutant is engineered to include at least one exogenous gene encoding a polypeptide that participates in the production of a lipid.

21. The method of claim 16 , wherein said algal mutant is cultured phototrophically.

22. The method of claim 21 , wherein said algal mutant is cultured in a pond or raceway.

23. The algal mutant of claim 11 , wherein said algal mutant has been genetically engineered by insertional mutagenesis, gene replacement, RNAi, antisense RNA, meganuclease genome engineering, one or more ribozymes, and/or a CRISPR/cas system.

24. The algal mutant of claim 1 , wherein said algal mutant is a diatom or Eustigmatophyte species.

25. The algal mutant of claim 1 , wherein said algal mutant is mutated in a gene encoding a polypeptide having at least 85% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 4 or the amino acid sequence of SEQ ID NO: 8, wherein said polypeptide recruits to pfam PF02135 and comprises an extended TAZ zinc finger domain comprising an amino acid sequence having at least 85% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 9 or the amino acid sequence of SEQ ID NO: 10.

26. The algal mutant of claim 25 , wherein said TAZ zinc finger domain comprises an amino acid sequence having at least 90% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 9 or the amino acid sequence of SEQ ID NO: 10.

27. The algal mutant of claim 1 , wherein said algal mutant is mutated in a gene encoding a polypeptide having at least 85% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 6 or the amino acid sequence of SEQ ID NO: 21, wherein said polypeptide recruits to pfam PF00249 and comprises an extended myb-like DNA-binding domain comprising an amino acid sequence having at least 85% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 22 or the amino acid sequence of SEQ ID NO: 23.

28. The algal mutant of claim 1 , wherein said extended myb-like DNA-binding domain comprises an amino acid sequence having at least 90% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 22 or the amino acid sequence of SEQ ID NO: 23.

29. The algal mutant of claim 1 , wherein said algal mutant is mutated in a gene encoding a polypeptide having at least 85% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 63 or the amino acid sequence of SEQ ID NO: 66, wherein said polypeptide comprises a domain comprising an amino acid sequence having at least 85% amino acid sequence identity to the amino acid sequence of SEQ ID NO: 64.

30. The algal mut

Assignees

• Synthetic Genomics Inc

Inventors

• Bailey Shaun
• Mccarren Jay
• Lieberman Soyan Leung
• Meuser Jonathan E
• Romano Anna E
• Yee Daniel
• Soriaga Leah
• Brown Robert C
• Schwartz Ariel S

Classifications

• C07K14/415

  from plants · CPC title

• C12N15/8269

  Photosynthesis · CPC title

• C12N15/825 Primary

  involving pigment biosynthesis · CPC title

• C12N1/36

  Adaptation or attenuation of cells · CPC title

• C12N15/01

  Preparation of mutants without inserting foreign genetic material therein; Screening processes therefor · CPC title