Comparative Transcriptomic Analysis Uncovers Genes Responsible For The DHA Enhancement In The Mutant Aurantiochytrium SP

DHA(Docosahexaenoic acid),known as “brain gold”,is a long-chain fatty acid containing six double bonds and belongs to the omega-3 series of polyunsaturated fatty acids.DHA cannot be synthesized by the human body so that it must be obtained from diets,and it plays a very important role in regulating and promoting various physiological activities of the human body.Aurantiochytrium,a marine fungus,is widely used in the DHA production due to its fast growth and high oil content.In this study,Aurantiochytrium sp.PKU # Mn16 was used as the wild-type.Ultraviolet irradiation(50 W,30 s)and quizalofop-p-ethyl(concentration 25 ? mol/L)were chosen as the breeding method of Thraustochytrid to obtain high yield of DHA.A total of 135 mutant mutant strains were obtained in the experiment.Among them,97 strains with higher biomass than wild type were cultured under the same conditions.According to the comparative analysis of biomass,fatty acid content,and DHA content,14 dominant mutant strains(named X1 to X14)were selected.Eight mutants(X1,X2,X3,X4,X5,X9,X11,X14)had increased oil production compared to the wild type,which were 1.09,1.79,1.26,1.25,1.23,1.35,1.08,1.64 times higher than the wild tpye,respectively.The DHA production of the four mutants(X2,X3,X5,X14)was significantly higher than that of the wild type,in which the DHA content of X2 polyunsaturated fatty acids increased from 40.55% to 50.19%,which was an increase of 23.77% compared to the wild type.Interestingly,9.07% of DPA was observed in mutant X2 instead of EPA.Ten generation-cultivation proved that the mutant X2 was genetically stable in the fatty acid production.Comparative transcriptome studies of wild-type and dominant mutagenic strains(X2)showed that there were 39826 differentially expressed genes(DEGs).In DEGs,1350 genes were down-regulated and 1945 genes were up-regulated.RNA-Seq analysis of differential expression and RT-q PCR studies showed that the m RNA expression levels of Co A transferase(Co AT),acyltransferase(AT),enoyl reductase(ER),dehydratase(DH)and methyltransferase(MT)were up-regulated in mutant X2,which is related to a significant increase in fatty acid synthesis in Aurantiochytrium sp.X2.In this research,we screened dominant strains of highyield polyunsaturated fatty acids,compared the transcriptomes of wild-type and dominant mutant strains,as well as obtained the key enzyme genes that affected the synthesis of DHA.This study provided a theoretical basis for the construction of DHA homologous recombination expression vectors.