Construction of shRNA of Fulminant Hepatitis Related Gene mfgl2 and Investigation of Its Biological Effects in vitro  

作　　者：Dong XI Zhi-Mo WANG Sui GAO Chuan-Long ZHU Jian-Wen GUO Xiao-Ping LUO Qin Ning 

机构地区：[1]Laboratory of Infectious Immunology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China

出　　处：《中国病毒学》2007年第5期366-373,共8页Virologica Sinica

基　　金：National Natural Science Foundation of China (No.30571643, No.30672380);National Key Basic Research Program of China (No. 2005CB2401,2007CB512904)

摘　　要：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.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.

关 键 词：爆发性肝炎 mfgl2基因 SHRNA RNA干涉 

分 类 号：Q78[生物学—分子生物学] R575.1[医药卫生—消化系统]