关键词: 过表达, NSPc1, 慢病毒载体, 多梳家族

Abstract: Objective To construct and identify lentiviral over-expression system to study the function of NSPc1 using over-expression technique．Methods The cDNA sequence of human NSPc1 was chosen to design the primer，and restricted enzyme site were added to primer sequences．The DNA purified from PCR were digested with double restriction enzymes and then linked with linearized pLenti6-TO-EGFP-TRIP．The recombinants were identified by restriction digestion and DNA sequencing．The plasmids of positive clones were transfected into 293T cells, and Western blot was used to identify the over-expression efficiency of the lentiviral vector. Then 293T cells were used to package the lentiviral particles and infected by obtained lentiviral particles. The over-expression efficiency of NSPc1 lentiviral system was examined by Western blot. Results The human NSPc1 sequences was successfully inserted into pLenti6-TO-EGFP-TRIP vector，which was identified by double restriction digestion and DNA sequencing．Western blot validated that both the lentiviral vector and lentiviral particle can effectively over-express human NSPc1 gene in 293T cells. Conclusion The construction of over-expression lentiviral system of human NSPc1 gene makes the further study of function of NSPc1 gene possible using over-expression technique．