Construction of SERCA Interference Lentivirus Expression Vector and Establishment of Stable PC 12 Cell Line / 中国药学杂志

ABSTRACT

OBJECTIVE: To construct the SERCA gene interference lentivirus expression vector and establish stable transfected PC 12 cell line. METHODS: The interference sequence targeting at rat SERCA gene was designed and synthesized. pGag/Pol, pRev, and pVSV-G were co-transfected into 293T cells. The lentivirus particles were packaged and generated. The virus titer was detected. PC 12 cells were transfected for establishing the stable cell line; RT-PCR and Western blot were used to detect SERCA gene and protein expression in stable PC 12 cells,and the RESULTS were compared with those in the control group. RESULTS: The lentivirus expression vector targeted at SERCA was successfully constructed and the virus titer was 3×108 U•mL-1. Stable transfected PC 12 cells line was established. The effective interference verification revealed that shSERCA could significantly reduce the mRNA and protein levels of SERCA (P<0.01). CONCLUSION: The shRNA lentiviral expression vector of SERCA gene is successfully constructed and the PC 12 cell line stably interfering with SERCA expresion is established.