Viral Hepatitis
Copyright ©2006 Baishideng Publishing Group Co., Limited. All rights reserved.
World J Gastroenterol. Feb 21, 2006; 12(7): 1038-1042
Published online Feb 21, 2006. doi: 10.3748/wjg.v12.i7.1038
Construction of eukaryotic expression plasmid containing HCV NS3 segment and protein expression in human HL-7702 hepatocytes
Jun-Wu Li, Ke Li, Jing Jiang, Xiao-Liang Xu, Ze-Qi Huang
Jun-Wu Li, Ke Li, Jing Jiang, Xiao-Liang Xu, Ze-Qi Huang, Microbiology and Immunology Department, Medical College, Jinan University, Guangzhou 510632, Guangdong Province, China
Supported by research fund from Ministry of Education of China for Studying Abroad, No. [2000]479; Natural Science Foundation of Guangdong Province, No. [2001]10-010371
Correspondence to: Dr. Jun-Wu Li, Microbiology and Immunology Department, Medical College, Jinan University, Guangzhou 510632, Guangdong Province, China. ljw6610@163.com
Telephone: +86-20-31308208; Fax: +86-20-85220730
Received: May 31, 2005
Revised: July 2, 2005
Accepted: August 20, 2005
Published online: February 21, 2006
Abstract

AIM: To construct the eukaryotic expression plasmid containing HCV NS3 segment and to analyze the expression of NS3 protein in normal human hepatocyte HL-7702.

METHODS: We amplified HCV NS3 fragment from plasmid pBRTM/HCV 1-3011 containing the whole length of HCV genome, recombined it with expression vector pcDNA3.1(-) to form the eukaryotic expression vector pcDNA3.1(-)/NS3, and transfected human HL-7702 hepatocytes with the recombined plasmid by cationic polymers. The expressed HCV NS3 protein was detected and analyzed by immunohistochemical method and Western blot.

RESULTS: The amplified NS3 fragments had correct molecule weight and sequence. The successfully constructed eukaryotic expression plasmids were transfected to HL-7702 cells. The expressed NS3 proteins had correct molecular weight 70000.

CONCLUSION: Eukaryotic expression vector pcDNA3.1 (-)/NS3 containing NS3 segment of HCV can be constructed, the sequence of NS3 fragments is consistent with the template. Normal human HL-7702 hepatocytes can efficiently express specific HCV NS3 protein in vitro.

Keywords: HCV; Structural protein; Expression