Construction and identification of mouse RelB siRNA-expressing lentiviral vectors

Scientific Research and Essays

Heng-cheng Zhu¹ 

关键词:  ;    siRNA-expressing lentiviral vector;    construction;    identification.;   

DOI  :  10.5897/SRE10.654

学科分类：社会科学、人文和艺术（综合）

来源: Academic Journals

【 摘 要 】

To determine the silencing efficiency of target genes by lentiviral vector-derived siRNA, this work constructed mouse RelB siRNA-expressing lentiviral vectors. Positive clones were screened by polymerase chain reaction (PCR) and identified by enzyme digestion as well as subsequent sequencing. RM-1 cells were transfected with liposome encapsulated siRNA, and Western blot and PCR were employed to detect the expressions of RelB in RM-1 cells followed by determination of the silencing efficiency. 293T cells were cotransfected with lentiviral vector and virus packaging plasmids mediated by calcium phosphate, and the virus titer was detected by measuring the GFP expression. PCR assay and sequencing demonstrated the RelB siRNA-expressing lentiviral vectors were successfully constructed. Real time- polymerase chain reaction (RT-PCR) and western blot showed the most effective target sequence of RelB was 5’-TGTCGTCAGGATCTGCTTC-3’. The lentivirus RNAi vector of relB which significantly inhibited relB expression was constructed successfully.

【 授权许可】

CC BY