Citation: YANG Zhen, JIANG Jiang—dong, QI Zi—bai, CHEN Hong—shan, Zhang Hua—yuan, LI He—ming. Cloning,Expression and Analysis of HCV Ns5b Polymerase gene from Chinese serum .VIROLOGICA SINICA, 2003, 18(6) : 530-533.

Cloning,Expression and Analysis of HCV Ns5b Polymerase gene from Chinese serum

  • Available online: 25 December 2003
  • Abstract:To ampUfy Hepatitis C virus(HCV)polymerase sequence and express HCV NS5b protein in order to,in the next,establish a molecular model of HCV replication.RT—PCR method was used to amplify HCV NS5b polymerase gene from a Chinese HCV(+1 patient’S serum. e polymerase fragment Was constructed in the plasmid pET一30a an d expressed in E coli.HCV NS5b protein Was identified by W lestern blot with a monoclonal an tibody against HCV NS5b.Sequence analysis showed tllat the isolate had 69% 一95% an d 89%一97% homologies to the reported HCV sequences in respect to nucleotide an d amino acid respectively.HCV NS5b gene was expressed at a high level an d the molecular weight ofthe expressed prod uct was 65kDa.which was within the ran ge of expectation.This protein Was confirmed to be HCV NS5b by Western blot.HCV NS5b from a Chinese patient Can be Success—fully cloned an d expressed with pET一30a plasmid an d E coli. is protein could be used to establish a molecular mod el of HCV replication.

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    Cloning,Expression and Analysis of HCV Ns5b Polymerase gene from Chinese serum

    • 1. National lnsititute for the Control of Pharmaceutical and Biology Products.Beijing 100050,China

    Abstract: Abstract:To ampUfy Hepatitis C virus(HCV)polymerase sequence and express HCV NS5b protein in order to,in the next,establish a molecular model of HCV replication.RT—PCR method was used to amplify HCV NS5b polymerase gene from a Chinese HCV(+1 patient’S serum. e polymerase fragment Was constructed in the plasmid pET一30a an d expressed in E coli.HCV NS5b protein Was identified by W lestern blot with a monoclonal an tibody against HCV NS5b.Sequence analysis showed tllat the isolate had 69% 一95% an d 89%一97% homologies to the reported HCV sequences in respect to nucleotide an d amino acid respectively.HCV NS5b gene was expressed at a high level an d the molecular weight ofthe expressed prod uct was 65kDa.which was within the ran ge of expectation.This protein Was confirmed to be HCV NS5b by Western blot.HCV NS5b from a Chinese patient Can be Success—fully cloned an d expressed with pET一30a plasmid an d E coli. is protein could be used to establish a molecular mod el of HCV replication.

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