The Adversity Reaction Of Diacylglycerol Acyltransferase Promoters In Chlamydmonas Reinhardtii

With the increasingly serious of environment problems such as atmospheric haze, The search for renewable energy is now a major research question for a variety of reasons, ranging from energy security to reducing carbon emissions. Microalgae has become a very advantage raw material of biodiesel with its rapid growth rates, occupy less and the high unit output, It has become a research hotspot of its fat metabolic pathway and how to increase oil production in recent years.In this paper, We studied promoters of CrDGAT1、 CrDGAT2-1、CrDGAT2-5, the genes of two key enzymes in the last step of fat metabolic pathway synthesis TAG in C. Reinhardtii. We First measured the biomass and fat content of C. Reinhardtii in nitrogen/sulfur/phosphorus deficiency stress conditions, results showed that lacking of nitrogen or sulfur or phosphorus is advantage to the oil accumulation, but reduce the biomass at the same time in Microalgae. Real time PCR results also indicated that the mRNA expression quantities of CrDGAT1、CrDGAT2-1、CrDGAT2-5in CC425cultured with HSM-P, low temperature and HSM-N were higher than CC425cultured with normal HSM medium.Tested the enzyme activity of reporter gene ARS by constructing deletion mutations of CrDGATl, CrDGAT2-1CrDGAT2-5promoter fragment introduced into C. Reinhardtii CC425. It showed that CrDGAT1promoter responded only to low temperature rather than other stress conditions; CrDGAT2-1promoter responsed to low temperature and HSM-P conditions, and had a weaker response to20μ M2,4-D、20μ M IAA、high salt (170mm NaCl)、HSM-K and HSM-Fe conditions; CrDGAT2-5had no response to the above conditions. A further study of CrDGAT2-1promoter and found that-319/-247regions are essential for inducible expression under the conditions of low temperature and HSM-P. The scanning mutagenesis analysis of these regions identified the cis-acting elements:low temperature at-312/-275; HSM-P at-319/-275and-261/-247.After the CrDGAT2-1P::ARS cloning vector inserted into CC425, the mRNA expression quantities of ARS and the endogenous genes appeared a higher expression under the conditions of low temperature、HSM-P and HSM-N, But under the condition of HSM-N, these three genes did not appear response, It may be associated with other epigenetic, and that still need further exploration.It had a preliminary analysis of CrDGAT1, CrDGAT2-1and CrDGAT2-5promoters reaction conditions in this paper, Which could provide theory basis for us to modify the promoters next steps. The mystery of promoter response needs an in-depth study.