Construction of shRNA of Fulminant Hepatitis Related Gene mfgl2 and Investigation of Its Biological Effects in vitro
- Author:
Dong, XI
;
Zhi-Mo, WANG
;
Sui, GAO
;
Chuan-Long, ZHU
;
Jian-Wen, GUO
;
Xiao-Ping, LUO
;
Qin, NING
- Publication Type:Journal Article
- Keywords:
Fulminant hepatitis;
fgl2 prothrombinase;
RNA interference;
shRNA
- From:Virologica Sinica
2007;22(5):366-373
- CountryChina
- Language:Chinese
-
Abstract:
This study was designed to explore the RNA interference technique in inhibition of the expression of the mouse fibrinogen like protein 2 (mfgl2), which has been reported to be involved in the development a variety of diseases including fulminant viral hepatitis. A plasmid named p-mfgl2shRNA,complementary to the sequence of mfgl2 was constructed, while another short hairpin RNA (shRNA)which was a mutated form of the mfgl2shRNA sequences was used as a control. A plasmid named pEGFP-mfgl2 expressing the mfgl2-EGFP fusion protein was also constructed for the screening of the effect of p-mfgl2shRNA on mfgl2 expression. By cotransfection of p-mfgl2shRNA and pEGFP-mfgl2 or pcDNA3.1-mfgl2 expression construct into CHO cells or HeLa cells, the inhibition of mfgl2 expression by mfgl2shRNA was analyzed by direct observation through fluorescent microscopy, FACS, RT-PCR and immunohistochemistry staining. The experiments showed the significant inhibitory effect of p-mfgl2shRNA on mfgl2 expression at 48h post-transfection in both CHO and Hela cell lines with the inhibitory efficiency as high as 80.1%. The study demonstrated that the construct of p-mfgl2shRNA successfully interfered with the mfgl2 expression in vitro.
