Construction of a GFP/Puro double-labeled lentiviral vector containing CK8 interfering RNA and its effect on cell apoptosis in vitro / 南方医科大学学报

ABSTRACT

<p><b>OBJECTIVE</b>To construct a GFP/Puro double-labeled lentiviral expression vector for CK8 silencing and assess the effects of CK8 silencing on cell apoptosis.</p><p><b>METHODS</b>The siRNA sequences of CK8 were inserted into the lentiviral expression vector GV248 and transfected into 293T cells with the packaging plasmids PMD and SPA. The lentivirus was collected at 24 and 36 h post-transfection. Flow cytometry was used to detect the virus titer and the positive cells were selected with puromycin. The knockdown of CK8 was examined by Western blotting. The effect of CK8 down-regulation on cell apoptosis induced by cisplatin was detected with Annexin V/PI staining.</p><p><b>RESULTS AND CONCLUSION</b>We successfully constructed CK8 interference lentiviral vector and obtained a stable cell line with CK8 knock-down that was sensitive to cisplatin-induced apoptosis.</p>