谷子糖基转移酶基因SiDDOST的克隆及表达载体的构建OA

In order to explore the biological function and regulatory role of glycosyltransferase gene in foxtail foxtail foxtail fox Mao Maoliang amplified the 1 338 bp cDNA sequence of the glycosyltransferase gene(SiDDOST)containing the complete coding region,and analyzed the characteristics of the full-length sequence of SiDDOST by bioinformatics methods,and the target gene fragment was double-digested with the restriction enzyme Nco I and Pml I,respectively,and the gene fragment and the vector fragment were recov-ered,and the glycosyltransferase gene fragment was linked to the overexpression vector pCAMBIA3301 by T4DNA ligase,and the re-combinant expression vector was transformed into E.coli DH5α.It was verified by electrophoresis detection of recombinant expression vector and empty vector,PCR of bacterial solution,and double enzyme digestion analysis.The results showed that the CDS region of glycosyltransferase gene was 1 323 bp and encoded 441 amino acids,the range of 30 amino acids at the N-terminus of DDOST pro-tein was poorly conserved,and there were three insertion/deletion mutations in DDOST protein compared with 8 other plant species.This indicated that the gene fragment of interest had been successfully inserted into the expression vector and the transcription direc-tion was correct.In conclusion,the overexpression vector of foxtail millet glycosyltransferase gene constructed in this study lays a foundation for subsequent genetic transformation and functional verification in Arabidopsis thaliana,rice or foxtail millet.