| The resources of the earth are not inexhaustible.The research and acquisition of safer green sustainable energy is an urgent problem to be solved in the world.Biofuels is a research hotspot in today's society.Algae are the most readily available,biomass-rich source of energy,and its metabolic by-products contribute to the medical,health,and beauty fieldsmetabolic by-products contribute to the medical,health,and beauty fields.In plants,the production of isoprene and other plastid isoprenoids mainly includes the cytosolic mevalonic acid(MVA)pathway and the cytoplast 2-C-methyl-D-erythritol-4-phosphate(MEP)pathway.Primary metabolites and secondary metabolites produced by the MEP pathway have important contributions to medicine,chemicals,food,and health,such as lipids and triacylglycerol(TAG).Previous studies have shown that the the first rate-limiting enzyme in the MEP pathway is 1-deoxy-D-xylulose-5-phosphate synthase(DXS),which is from plastids,controls the synthesis of isopentenyl pyrophosphate(IPP),an isoprenoid precursor,to regulate the amount of primary metabolites and secondary metabolites in plants.If we strengthen the effect of this link,it is expected to increase the MEP pathway and increase the synthesis of oils such as TAG,thereby increasing the yield of terminal metabolites.In this study,Nannochloropsis ocenica was used as the research object.After successfully introducing the Arabidopsis thaliana DXS gene,the DXS gene overexpressing Nannochloropsis ocenica(Arabidopsis thaliana DXS overexpression Nannochloropsis ocenica,DXSoe)was obtained,and its growth and stress resistance were studied.And synthesis of related metabolites.The transformants DXSoel,DXSoe2 and DXSoe3 with excellent growth traits were obtained by comparison of growth rates under PCR and normal culture conditions.After using the DXS inhibitor clomazone(CLO,the foreign trade name is chlorhexidine),the transformants showed stronger stress resistance and growth performance than the wild type(1-3 times).The growth,stress resistance,biomass accumulation of wild plants and DXSoe1,DXSoe2 and DXSoe3 were compared by environmental factors(different illuminance,nitrogen content,trace element content)by spectrophotometry,microscopic counting,dry weight method and phenol sulfuric acid method.The effects of chlorophyll and total carbohydrate content.The results showed that the number of cells,chlorophyll content and total carbon content of transformants were 1-3 times higher than that of wild type,and had better physiological advantages(non-repetitive two-way analysis of variance,P<0.01 or P<0.05).In addition,we also determined the content of oils and triacylglycerols in marine microspheres.The oil content and TAG content(thin layer chromatography)of overexpressing transformants were 1-2 times higher than wild type(non-repetitive two-way analysis of variance,P<0.01 or P<0.05)(Parallel experiments were performed on all the above indicators,and the parallelism was good).It can be seen that the overexpression of DXS in the biomass of Nannochloropsis ocenica is feasible,which will provide an important basis and reference for the optimization of microalgae core metabolic pathways by metabolic engineering and synthetic biology,and provide reliable research foundation and reference for other related fields such as food,feed,etc.,which can provide a theoretical basis for the further application of engineering cells,more effective in solving energy shortages and environmental pollution problems. |
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