In Vitro Transcription of HCV 5 Non-Coding Region cDNA

epatitis C virus(HCV)5'non-coding region(5'NCR)cDNA with 302 bp obtained byreverse transcription polymerase chain reaction(RT-PCR)from the serum of a patient withchronic hepatitis C in Guangdong.Province was subsequently filled in recessed ends,purified andinserted into pUC19 plasmid vector.The recombinant plasmid pUN was sequenced.The targetgene in pUN was subcloned into EcoR I/Pst I sites of pSPORT I transcription vector.Alter therecombinant pSN was linearized,the transcription reaction in vitro was performed using SP6RNA polymerase or T_7 RNA polymerase。The sense RNA with 429 bp and anti-sense RNA with362 bp synthesized by SP6 RNA polymerase and T7 RNA polymerase respectively were identifiedby electrophoresis on agarose gel and by RT-PCR using HCV-specific primers.It was also ver-ified that the RNAs were transcripted from HCV5'NCR cDNA.The 5'NCR cDNA clone con-structed and RNA synthesized can be used as effective positive control templat for PCR and RTrespectively,which will be helpful in eliminating false