Cloning and expression of NS3 cDNA fragment of HCV genome of Hebei isolate in E.coli.

ABSTRACT

AIM:To obtain greater antigenicity of HCV NS3 protein.METHODS:The HCV NS3 cDNA fragment was amplified by reverse transcription polymerase chain reaction from the sera of the HCV infected patients.The DNA sequence was determined by dideoxy-mediated chain termination method using T7 polymerase.HCV NS3 protein was expressed in E. coli.RESULTS:Sequence analysis indicated that the HCV isolate of this study belongs to HCV-II; SDS-PAGE demonstrated an M(r) 23800 and an M(r) 22000 recombinant protein band which amount to 14% and 11% of the total bacterial proteins separately.Western blotting and ELISA showed NS3 protein possessed greater antigenicity.CONCLUSION:Recombinant HCV NS3 protein was expressed successfully, which provided the basis for developing HCV diagnostic reagents.