Cloning And Genetic Transformation Of Medicago Sativa OPT1 Gene

Oligopeptide transporter protein is a kind of important transporter in plants. Oligopeptide transporter has an important role in resistance to biotic and abiotic stress and development of plants. In this study, MsOPT1 gene was cloned from Medicago sativa, and then linked into the expression vector of pCBM, genetic transformed into Nicotiana tabacum and Arabidopsis thaliana for the research of gene function.The main findings are as follows:1. MsOPT1 gene cloning and sequence analysisAccording to amino acid sequence of AtOPT gene in Arabidopsis thaliana and blast in the Medicago truncatula genome database, find homologous sequence and design the primers, cloned a gene and named it MsOPT1. The MsOPT1 gene has a total of 411 nucleotides encoding 136 amino acids, no intron and one OPT domain.2. pCBM-MsOPT1 vector constructionConstruct the plant expression vector pCBM-MsOPT1, carrying with selection marker gene Bar and CaMV35 S promoter.3. MsOPT1 gene transformation into tobaccoUsing the method of agrobacterium mediated transformation, MsOPT1 gene was transformed into Nicotiana tabacum. Through resistance screening, PCR and RT-PCR, obtained positive transgenic tobacco plants of MsOPT1 gene.4. The phenotype analysis of transgenic MsOPT1 tobaccoThe results of qRT-PCR showed that the expression of transgenic plants were higher. Compared to the WT, transgenic MsOPT1 tobacco showed phenotype changes. The transgenic MsOPT1 tobacco has more content of Chlorophyll and Fe than those of the WT. Under iron stress, the seed germination rate of transgenic tobacco were higher than that of WT. The root of transgenic tobacco were longer than that of WT.5. MsOPT1 gene transformation into Arabidopsis thalianaThrough the inflorescence infection method, MsOPT1 gene was transformed into Arabidopsis thaliana. T0 and T1 plants were obtained.