Establishment of a Bladder Cancer Cell Line with Stable Knockdown of MEX3A Gene via Lentiviral-mediated Interference
10.12007/j.issn.0258-4646.2017.12.001
- VernacularTitle:慢病毒介导的靶向干扰MEX3A基因膀胱癌稳定细胞株的建立
- Author:
Chao FANG
1
;
Jing YIN
;
Qiushuang YU
;
Yunyun CHEN
;
Ying HUANG
Author Information
1. 中国医科大学附属盛京医院超声科
- Keywords:
MEX3A gene;
bladder cancer;
lentivirus;
cell line
- From:
Journal of China Medical University
2017;46(12):1057-1061,1066
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct an shRNA lentiviral vector targeting the MEX3A gene and establish a bladder cancer cell line with stable MEX3A gene knockdown.Methods Real-time PCR was performed to detect the MEX3A gene expression.The recombinant lentiviral vector targeting the MEX3A gene was constructed using the GV115 plasmid.After identification and sequencing,the vectors were co-transfected with the packaging vector into 293T cells to produce lentiviral particles,which were then transduced into bladder cancer cells after viral titer determination.The cell line stably expressing the siRNA was established by antibiotic selection,and real-time PCR was carried out to detect the efficiency of the knockdown.Results Both bladder cancer cell lines,5637 and T24,expressed the MEX3A gene,and its expression was higher in 5637 than in T24.The identification and sequencing results showed that the MEX3A-shRNA lentiviral vector was successfully constructed,and the virus titer was observed to be higher after packaging.The results of the real-time PCR showed that MEX3A gene expression was stably inhibited in 5637 cells after lentiviral transduction.Conclusion Lentivirus-mediated RNAi technology could successfully establish a cell line with stable MEX3A gene knockdown.
