Construction of Sirtl shRNA interfering vector and its effects on cell proliferation and apoptosis.
- Author:
Wenting DU
1
;
Sichong REN
;
Qifeng SUO
;
Mingming YANG
;
Dan HE
;
Ji LIU
Author Information
1. Department of Biochemistry and Molecular Biology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
- Publication Type:Journal Article
- MeSH:
Apoptosis;
genetics;
Cell Line, Tumor;
Cell Proliferation;
Genetic Vectors;
Humans;
RNA Interference;
RNA, Small Interfering;
genetics;
Sirtuin 1;
genetics;
Transfection
- From:
Journal of Biomedical Engineering
2011;28(5):972-975
- CountryChina
- Language:Chinese
-
Abstract:
This study was aimed to construct Sirt1 shRNA interfering vector and to analyze the effects of Sirtl on cell proliferation and apoptosis in HepG2, A549 and 293T cell lines. To design and synthisize Sirtl shRNA sequence then recombinate it to pGenesil-1.0 plasmid, the positive pGenesil-1.0-Sirtl vector clone was screened by effective detections and sequencing. The vectors were transfected into HepG2, A549, 293T cell lines, and Sirtl expression levels in these clones were detected by RT-PCR and Western-blot. These clone cell proliferation activities were detected by MTT, and these cells apoptosis incidences were detected by MTT experiment after treated with DOX. The results showed that Sirt1 shRNA interfering vectors were successfully screened. The levels of Sirtl expression in HepG2-sh, A549-sh and 293T-sh cells were significantly reduced compared with their control cells. It was indicated that the proliferation activities of these cells were impaired and anti-apoptosis capabilities of HepG2-sh, A549-sh and 293T-sh were also impaired notably. Sirt1 took an important role in maintaining cell proliferation and resisting cell apoptosis caused by DNA damage, and this result also provided theoretical information for the further research.
