Overexpression of acetyl-CoA carboxylase increases fatty acid production in the green alga Chlamydomonas reinhardtii
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Chlamydomonas reinhardtii is a photosynthetic unicellular model algae with multiple biotechnological advantages, and its fatty acids can be used to produce biofuels. Numerous studies suggest that acetyl-coA carboxylase (ACCa) catalyzes the first committed and rate-limiting step of fatty acid biosynthesis, thereby playing a central role in oil accumulation. Here, we cloned and overexpressed ACCa in C. reinhardtii to directly evaluate its effect on fatty acid synthesis. GC–MS analysis found that the unsaturated FAs contents of the CW15-24 and CW15-85 strains were 55.45% and 56.15%, which were significantly enriched compared to the wild type CW15 (48.39%). Under the optimized conditions, the content of lipid by overexpressed the ACCa gene in the mutant CW15-85 (0.46 g/l) was 1.16-fold greater than control through optimization of N and P sources. Altogether, our data clearly demonstrate that ACCa overexpression in C. reinhardtii can directly increase the synthesis of fatty acids.
KeywordsAcetyl-CoA carboxylase Biofuel Chlamydomonas reinhardtii Fatty acids Lipid
We would like to thank the native English speaking scientists of Elixigen Company (Huntington Beach, California) for editing our manuscript. This work was supported by the Natural Science Foundation of Fujian Province, China (Grant Number 2017J01622) and the Sugar Crop Research System (Grant Number CARS-170501).
Supplementary Fig. 1—Characterization of the PHKA overexpression construct by PCR detection of Acca and mCherry and restriction analysis. (A) Acca amplification from three independent PHKA clones confirms insertion of Acca coding sequence (lane M, molecular weight marker; lanes 1-3, Acca). (B) Restriction validation of PHKA clones (lane M, molecular weight marker; lane 1, products of PHKA; lanes 2-3, EcoR I digestion; lanes 4-5, EcoR V and EcoR I digestion). (C) mCherry amplification from C. reinhardtii mutant genomic DNA (lane M, molecular weight marker; lanes 1-5, mCherry).
Supplementary Fig. 2−Protein spot hybridization using anti-mCherry and anti-GAPDH antibodies. (A) GAPDH (endogenous control) is detected in wild type (CW15) and ACCa-overexpressing transgenic clones (CW15-24, CW15-85). (B) mCherry is detected only in the ACCa-overexpressing (CW15-24, CW15-85) clones, suggesting successful transfection of the PHKA overexpression vector.
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