Construction of a GFP/Puro double-labeled lentiviral vector containing CK8 interfering RNA and its effect on cell apoptosis in vitro
10.3969/j.issn.1673-4254.2013.12.11
- VernacularTitle:细胞角蛋白8 GFP/Puro双标干涉慢病毒系统的构建及对细胞凋亡的影响
- Author:
Yanchao JIN
1
;
Ronghua YIN
;
Weiwei ZHENG
;
Xiangzhen KONG
;
Yiqun ZHAN
;
Xiaoming YANG
;
Changyan LI
Author Information
1. 天津大学制药工程系
- Keywords:
CK8;
GFP/Puro double-marker lentiviral vector;
RNA interference;
apoptosis;
cisplatin
- From:
Journal of Southern Medical University
2013;(12):1761-1765
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct a GFP/Puro double-labeled lentiviral expression vector for CK8 silencing and assess the effects of CK8 silencing on cell apoptosis. Methods 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. Results and Conclusion We successfully constructed CK8 interference lentiviral vector and obtained a stable cell line with CK8 knock-down that was sensitive to cisplatin-induced apoptosis.
