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Volume 19 Issue 4
August 2004
    Citation: YANG Xiao-jun, YE Lin-bai*, GAO Jin-rong, LIU Jing, YANG Fan, YE Li, SHE Ying-long, LIAO Qing-jiao, WU Zheng-hui, ZHENG Yi. Stable Expression of NS5A Gene of HCV in Hela Cells and Inhibition to the Growth of Hela Cells .VIROLOGICA SINICA, 2004, 19(4) : 340-344.
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    Stable Expression of NS5A Gene of HCV in Hela Cells and Inhibition to the Growth of Hela Cells

    • 1.

      Institute of Virology, Wuhan University, Wuhan 430072, China

    • Corresponding author: YE Li, 
    • Available online: 25 August 2004
    • Full-length NS5A gene of Hepatitis C virus（HCV） was amplified by PCR, using the plasmid pBRTM/HCV 1-3011 containing HCV full-length open reading frame（ORF）as template, and cloned into the eukaryotic expressing plasmid pcDNA3.1（-）by DNA recombination technique. The recombin- ant vector was identified by digestion with restriction enzymes and polymerase chain reaction and by directly sequencing. Then both the recombinant vector pcDNA3.1（-）-NS5A and the control vector pcDNA3.1（-）were transfected Hela cells using LipoVecTM. The cells expressing NS5A stablely were selected by G-418 and further proved by RT-PCR and Western blot analysis. We found the growth of Hela cells expressing NS5A was slower than the cells transfected by pcDNA3.1（-）in the same culture condition, and the population doubling time of Hela cells expressing NS5A gene is increased about 50%(about 35-35 hours). There was no significant difference between the control cells and the cells transfected with pcDNA3.1(-) (about 23-24 hours). The results indicate that NS5A can inhibit the proliferation of Hela cells.
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      Stable Expression of NS5A Gene of HCV in Hela Cells and Inhibition to the Growth of Hela Cells

        Corresponding author: YE Li,
      • 1. Institute of Virology, Wuhan University, Wuhan 430072, China

      Abstract: Full-length NS5A gene of Hepatitis C virus（HCV） was amplified by PCR, using the plasmid pBRTM/HCV 1-3011 containing HCV full-length open reading frame（ORF）as template, and cloned into the eukaryotic expressing plasmid pcDNA3.1（-）by DNA recombination technique. The recombin- ant vector was identified by digestion with restriction enzymes and polymerase chain reaction and by directly sequencing. Then both the recombinant vector pcDNA3.1（-）-NS5A and the control vector pcDNA3.1（-）were transfected Hela cells using LipoVecTM. The cells expressing NS5A stablely were selected by G-418 and further proved by RT-PCR and Western blot analysis. We found the growth of Hela cells expressing NS5A was slower than the cells transfected by pcDNA3.1（-）in the same culture condition, and the population doubling time of Hela cells expressing NS5A gene is increased about 50%(about 35-35 hours). There was no significant difference between the control cells and the cells transfected with pcDNA3.1(-) (about 23-24 hours). The results indicate that NS5A can inhibit the proliferation of Hela cells.

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