Viral Hepatitis
Copyright ©2005 Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Jan 28, 2005; 11(4): 508-510
Published online Jan 28, 2005. doi: 10.3748/wjg.v11.i4.508
Detection of hepatitis B virus DNA by real-time PCR using TaqMan-MGB probe technology
Jin-Rong Zhao, Yu-Jie Bai, Qing-Hua Zhang, Yan Wan, Ding Li, Xiao-Jun Yan
Jin-Rong Zhao, Yu-Jie Bai, Qing-Hua Zhang, Yan Wan, Ding Li, Xiao-Jun Yan, Institute of Genetic Diagnosis, Fourth Military Medical University, Xi’an 710032, Shaanxi Province, China
Author contributions: All authors contributed equally to the work.
Correspondence to: Dr. Jin-Rong Zhao, Institute of Genetic Diagnosis, Fourth Military Medical University, 17 Changle West Road, Xi’an 710032, Shaanxi Province, China. zhaojrr@fmmu.edu.cn
Telephone: +86-29-83374772 Fax: +86-29-83285729
Received: December 10, 2003
Revised: December 13, 2003
Accepted: January 8, 2004
Published online: January 28, 2005
Abstract

AIM: To develop a real-time PCR for detecting hepatitis B virus (HBV) DNA based on TaqMan technology using a new MGB probe.

METHODS: Plasmid containing the sequence of X gene (1414-1744 nt) was constructed as HBV-DNA standard for quantitative analysis. A TaqMan-MGB probe between primers for amplification was designed to detect PCR products. The interested sequence contained in the plasmid and in clinical specimens was quantitatively measured.

RESULTS: The detection limit of the assay for HBV DNA was 1 genome equivalent per reaction. A linear standard curve was obtained between 100 and 109 DNA copies/reaction (r>0.990). None of the negative control samples showed false-positive reactions in duplicate. HBV DNA was detected in 100% (50/50) of HBV patients with HbeAg, and in 72.0% (36/50) with HBsAg, HBeAb and HBcAb. The coefficient of variation for both intra- and inter-experimental variability demonstrated high reproducibility and accuracy.

CONCLUSION: Real-time PCR based on TaqMan-MGB probe technology is an excellent method for detection of HBV DNA.

Keywords: Hepatitis B Virus; DNA; TaqMan-MGB probe; Real-time PCR