Expression of RNase H of human hepatitis B virus polymerase in Escherichia coli

• Published in: World journal of gastroenterology : WJG Vol. 9; no. 3; pp. 513 - 515
• Main Authors: Cheng, Hong, Zhang, Hui-Zhong, Shen, Wan-An, Liu, Yan-Fang, Ma, Fu-Cheng
• Format: Journal Article
• Language: English
• Published: United States Department of Pathology, Xijing Hospital, Fourth Military Medical University, Xi'an 710033, Shaanxi Province, China%Orthopedics Oncology Institute of Chinese PLA, Fourth Military Medical University, Tangdu Hospital, Xi'an 710038, Shaanxi Province, China 01-03-2003; Baishideng Publishing Group Inc
• Subjects: DNA-Directed DNA Polymerase; Escherichia coli - metabolism; Gene Transfer Techniques; Hepatitis B virus - metabolism; Humans; Ribonuclease H - genetics; Ribonuclease H - metabolism; Viral Hepatitis
• ISSN: 1007-9327; 2219-2840
• DOI: 10.3748/wjg.v9.i3.513

To amplify HBV-RNase H gene fragment and expression of RNase H for further use in the studies of HBV associated liver diseases.
The encoding gene of HBV-RNase H was separately amplified for the first half and second half (H1 and H2) by PCR from full length HBV gene and cloned into pT7Blue-T vector. Clones were first screened by digestion with XbaI and Hind III enzyme for the correct size, and analyzed further by DNA sequencing. The RNase H1 and H2 fragments isolated from XbaI and Hind III digestion products of pT7 Blue-RNase H plasmid were ligated to the GSTag expressing vectors separately, and expressed in E.coli BL21. The expressed proteins were checked by PAGE gel and Western blot.
Both H1 and H2 nucleotide seqences consisting of known genes and proteins, in correct size, were further confirmed by Western blot to be the GST and RNase H1 or H2 fusion proteins.
The successful cloning and expression of HBV-RNase H will contribute to further research and application in HBV-associated diseases.