Breeding And Optimization Of DHA-producing Strain Of Schizochytrium Sp.Mutagenesis Based On ARTP

Docosahexaenoic acid(DHA)is a type of omega-3 polyunsaturated fatty acidwith significant impacts on human health.It serves as supplementary nutrient for functional food and milk powder for cancer prevention and treatment,hypertension and cardiovascular diseases.Schizochytrium sp.is regarded as an ideal marine protist for DHA production to achieve fatty acid biosynthetic capacity,easy manipulation,rapid growth,harmless to human body and other advantages.However,the productivity of DHA in Schizochytrium needs to be further improved.The selection and fermentation conditions of high DHA producing Schizochytrium species were studied in detail in this paper.In order to obtain high-yield DHA mutant strains of Schizochytrium sp.PKU Mn4,two rounds of mutagenesis were carried out with ARTP mutagene instrument.After screening with ketene,vanillin phosphate and shaking flask fermentation,two high-yield DHA mutant strains A78 and A92 were obtained.The yield of DHA was0.94 g/L and 0.87 g/L,respectively,which were 54% and 42% higher than that of wild-type strains.In order to further increase DHA production,the metabolic pathway of fatty acid synthesis was manipulated.Sodium citrate,citric acid and biotin were added as alternative carbon sources respectively.The optimal results of orthogonal test showed that the DHA production in mutants A78 and A92 were 1.29 g/L and 1.21g/L,respectively,which were 51% and 47% higher than that of the control group.PUFA yields were 1.56 g/L and 1.45 g/L,respectively,which were 29% and 32%higher than that of the control group.DHA yield increased by 110% and 98%compared with wild-type strains.Secondly,the composition of mutant fermentation medium was optimized.Through orthogonal test,DHA yield of mutant A78 and A92 was 1.65 g/L and 1.44 g/L,respectively,which was 85% and 69% higher than that of the control group,1.85 g/L and 1.61 g/L higher than that of the control group,53%and 46% higher than that of the control group,and 170% and 136% higher than that of the wild-type strains.In summary,high-yield DHA mutant was obtained and screened by ARTP mutation.After adding exogenous additives and optimizing fermentation conditions,the production of DHA was significantly improved.It verified the feasibility of breeding method of combining ARTP mutation and enoxazone screening to obtain high-yield DHA producing Schizochytrichum mutant,and provided new germplasm for industrial production of DHA.Finally,the results showed that the DCW,DHA and PUFAs of mutant A78 increased by 49%,205% and 135% compared with wild type,and the DCW,DHA and PUFAs of mutant A92 increased by 42%,165% and 121% compared with wild type.