Mutation-screening Of Mortierella Isabellina With High-yield Lipid And Optimization Of Fermentation Medium

Lipid is not only the basic substances of composing and maintaining life, and it isa kind of important industrial raw material. With the increasing population andcontinued demand for industrial utilization, the plant and animal sources alone cannotmeet the total requirement. Therefore, the development and utilization of new oilsources——microbial oils, have important theoretical and practical application value.Usually, the fatty acid composition of microbial lipid is similar to that of plantoils. And it contains high fractions of polyunsaturated fatty acids （PUFA）, such asarachidonic acid （AA）, γ-linolenic acid （GLA）, eicosapentaenoic acid （EPA）,docosahexenoic acid （DHA） and so on. These PUFAS with special function have moreand more vital role in promoting the human health aspect. Now, production ofmicrobial lipid cost higher than plant lipid, so the study of microbial lipid mainlyconcentrates in the production of high economic value nutritional oils and specialindustrial oils.Compared with the animal and plant oil, microbial lipid has many advantages,such as such as short life cycle, less labor required, less affection by venue, season andclimate, plentiful and cheap raw material. However, microbial lipid also has itslimitation, and the most obvious aspect is the lower yield of lipid and higher cost. Inorder to improve yield and achieve its industrial application, this article has done theresearch work as follows:1. In order to obtain lipid producing strain with high-yield, the strain MortierellaIsabellina As3.3410was treated by ultraviolet, diethyl sulfate, microwave. It wasfound that the optimum ultraviolet time was20s, the diethyl sulfate at20min with adoses of0.8%was optimum, the best time of intermittent microwave was40s.Comparison of three kinds of mutagenesis method, the ultraviolet and diethyl sulfatewere optimum mutagenic methods for Mortierella Isabellina.2. To establish a high-throughput and easy screening method for mutant strains, the curelenin （a fatty acid synthase inhibitor） was tried to screening of MortierellaIsabellina. The results showed that the8.96×10^-6mol/L curelenin was the bestconcentration for screening mutant strain with high-yield. The lipid yield wasmeasured with acid-heating extraction, and the accuracy of curelenin resistancescreening was verified. After repeated mutation-screening, the strain Z10was selectedwith high-yield and lipid yield was10.76g/L, increasing by73%than original strain.The production characteristic of Z10had no significant change through successivetransfer of culture. And it was found that genetic characteristics of Z10were ratherstable. Additionally, the fermentation characteristic of high-yield mutation strain Z10was studied.3. Additionally, the surface response method was used to optimize fermentationmedium. The four significant factors （glucose, yeast extract, MgSO₄, KH₂PO₄） wereoptimized using response surface methodology （RSM）, and the optimized parametersof fermentation medium were as follows: glucose90.29g/L, yeast extract4.14g/L,MgSO40.96g/L, KH2PO41.99g/L. Under the optimized condition, the lipid yield ofMortierella Isabellina reached as high as12.01g/L, and the average lipid yield up to12.19g/L in the test, increased by13.29%.