Genetic engineering of microalgae for enhanced lipid production

Abstract

Genetic transformation of microalgae remains a challenge due to poor intracellular delivery of exogenous molecules. This limitation is caused by the structure and composition of the cell wall and cell membrane of each species.Moreover, successful delivery of proteins or nucleic acids cannot be assessed by determining transformability since their functionality is not always known in the studied microorganism. We propose a quick and effective screening tool for the prediction and optimization of electroporation settings by monitoring cell permeability and viability using Sytox Green and propidium iodide respectively.We determined voltage settings for the microalgae Chlamydomonas reinhardtii, Chlorella vulgaris, Neochloris oleoabundans and Acutodesmus obliquus. To evaluate the predicted settings, we delivered labelled DNA and proteins into the cells. We demonstrated that high transformation efficiencies can be accomplished when predicted values were applied with functional plasmids.Additionally, we increased transformation efficiencies by testing cell concentrations, light intensities and fragment sizes. This method can be used to determine suitable transformation conditions for non-transformed microalgae species and to increase the insight on established transformation protocols.

Strategic Development of Aurantiochytrium sp. Mutants With Superior Oxidative Stress Tolerance and Glucose-6-Phosphate Dehydrogenase Activity for Enhanced DHA Production Through Plasma Mutagenesis Coupled With Chemical Screening

“…At present, the advancement in microbial genetic engineering and synthetic biology has led to a significant increase in super strain development for the production of important metabolites of interest, including PUFAs from the microalgae ( 17 , 42 , 43 ). Nevertheless, genetic engineering technology in thraustochytrids including the Aurantiochytrium sp.…”

Carotenoid Production from Microalgae: Biosynthesis, Salinity Responses and Novel Biotechnologies

“…Targeted genetic engineering could generate specific insertions, deletions or substitutions in the host genome while avoiding random results [113]. Up until now, a lot of genes related to salt responses have been recognized, such as nhaP encoding a Na + /H + antiporter, codA encoding choline oxidase to synthesize compatible solutes and Dps encoding DNA-binding proteins (Dps).…”

Increased Lipids in Chlamydomonas reinhardtii by Multiple Regulations of DOF, LACS2, and CIS1

“…C. reinhardtii , a model photoautotrophic microalga, is a promising material for transgenic engineering because of its high photosynthetic efficiency, fast growth rate and diverse transformation systems [ 4 ]. Lipid engineering for higher production has been widely studied in C. reinhardtii regarding FA synthesis, Kennedy pathway, transcription factors, and NADPH generation [ 5 ]. FA synthesis, the first step of lipid formation, occurs in the chloroplast of C. reinhardtii using acetyl-CoA as a substrate.…”

“…FA synthesis, the first step of lipid formation, occurs in the chloroplast of C. reinhardtii using acetyl-CoA as a substrate. Acetyl-CoA carboxylase (ACC), malonyl-CoA ACP transacylase (MAT), 3-ketoacyl-ACP synthase (KAS), 3-ketoacyl-ACP reductase (KAR), 3-hydroxyacyl-ACP dehydrase (HD), and enoyl-ACP reductase (ENR) are the key enzymes involved in this process [ 5 ]. Glycerolipids are synthesized stepwise in the Kennedy pathway through the catalysis of glycerol 3-phosphate acyltransferase (GPAT), lysophophatidic acid acyltransferase (LPAT), phosphatidic acid phosphatase (PAP), and diacylglycerol acyltransferase (DGAT).…”