Citation: TIAN Jing, JU Gui-Beng, FANG Qi. cDNA Synthesis,Cloning and Partial Nucleotide Sequence of Grass Carp Hemorrhage Virus(Gcnv) .VIROLOGICA SINICA, 1999, 14(1) : 87-92.

cDNA Synthesis,Cloning and Partial Nucleotide Sequence of Grass Carp Hemorrhage Virus(Gcnv)

  • Available online: 05 March 1999
  • Grass Carp Hemorrhage Virus (GCHV) was purified from infected fish cell culture by using different centrifugation. Its total genome was isolated by proteinase K treatment following phenol/chloroform extraction. The individual segment of GCHV dsRNA was separated by running 1% agarose gel electrophoresis and recovered by Gel Extraction Kit. Each segment was denatured in 90% DMSO by heating at 70℃ for 15min. The cDNAs of several segments were synthesized with random hexamers method and cloned into EcoRV site of pZErO 2.0 vector, then transformed into TOP10 E.coli competent cell by electroporation transformation. It was showed that the recombinant plasmids contained GCHV RNA genome of interest by dot hybridization besides both restriction analysis and PCR amplification. The partial DNA sequence of GCHV segment 11 was analysed.

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    cDNA Synthesis,Cloning and Partial Nucleotide Sequence of Grass Carp Hemorrhage Virus(Gcnv)

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    Abstract: Grass Carp Hemorrhage Virus (GCHV) was purified from infected fish cell culture by using different centrifugation. Its total genome was isolated by proteinase K treatment following phenol/chloroform extraction. The individual segment of GCHV dsRNA was separated by running 1% agarose gel electrophoresis and recovered by Gel Extraction Kit. Each segment was denatured in 90% DMSO by heating at 70℃ for 15min. The cDNAs of several segments were synthesized with random hexamers method and cloned into EcoRV site of pZErO 2.0 vector, then transformed into TOP10 E.coli competent cell by electroporation transformation. It was showed that the recombinant plasmids contained GCHV RNA genome of interest by dot hybridization besides both restriction analysis and PCR amplification. The partial DNA sequence of GCHV segment 11 was analysed.

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