Physiological And Molecular Mechanisms Of BrDMC1 Response To Salt Stress In Brassica Rapa

DMC1 protein is a typical member of the recombinase family of eukaryotes,with homology search for single-stranded DNA to achieve the biological function of double-strand break repair.It plays an important role in repairing DNA oxidative damage caused by stress.Soil salinization has become an extremely important agricultural problem in the world,so identifying genes that respond to salt stress is of great significance for enhance plant stress tolerance.In this study,we discovered that Br DMC1 is involved in the DNA repair process through bioinformatics,and may also be involved in the response of plants to stress.In this experiment,RNA interference technology was used to down-regulate the expression level of Br DMC1 gene in B.rapa.Analysis The difference in stress resistance to further explore the possible mechanism of Br DMC1 in response to salt stress.The results of this study are as follows:1.There are two copies of DMC1 in named Br DMC1.A01 and Br DMC1.A03 in B.rapa.Analysis of its functional domain shows that Br DMC1 protein has a typical recombinase Rec A domain and is highly conservative in function.It was verified by RT-PCR that Br DMC1 functions as a single copy,and Br DMC1.A03 was not transcribed.2.We examined transcriptional level of Br DMC1 under different stress treatments such as Na Cl,Mannitol and ABA,the results showed that Br DMC1 was highly induced by salt stress.It indicates that Br DMC1 is high likely to participate in the response to salt stress.3.The Br DMC1 gene is expressed in the roots,stems,leaves,flower and seed of Brassica rapa,with the highest expression in flowers,followed by seeds.GUS staining results showed that GUS signal could be detected in flower and germinated seeds.Subcellular localization results show that Br DMC1 protein is located on the nucleus.4.Cloning and analysis the Br DMC1 promoter,and predicting the cis-acting element of the Br DMC1 promoter and finding that it contains stress response element.The expression of Br DMC1 promoter under stress treatment was verified by transient expression of GUS in tobacco leaves.The results showed that Br DMC1 promoter was induced by these stresses.5.Created RNAi-Br DMC1 transgenic plants of B.rapa and inhibited the expression of Br DMC1 gene.It was found that the germination rate of RNAi-Br DMC1 seeds was significantly reduced compared with the wild type under salt stress treatment.No difference in germination rate while under normal(control)treatment.Further measurement of the antioxidant enzyme activity and osmotic protective substance content involved in the stress response revealed that the antioxidant enzyme activity in RNAi-Br DMC1 seeds decreased and the soluble sugar content decreased.the ability of ROS scavenging is reduced,reactive oxygen is accumulated in large amounts,the level of MDA content is increased,and membrane oxidation is increased.A large amount of accumulated reactive oxygen species leads to increased DNA oxidative damage.Result a serious lag germination in RNAi-Br DMC1 seed.These results further proved that Br DMC1 may participate in the adaptation of plant to salt stress tolerance.