Detection for genotypes of hepatitis B virus by nested PCR with multiplex pairs of primers

Objective To determine the genotypes of hepatitis B virus (HBV) by nested PCR with multiplex pairs of genotype-specific primers.Methods Ten outer and inner primers were designed on the basis of the conserved nature of nuleotide sequences in regions of the Pre-S1 through S genes,in which 8 inner primers were devided into mix A and B to amplify HBV of genotype A,B,C and D,E,F respectively.The two different products from one sample in second-round PCR were separately electrophoresed on a 3% agarose gel.Genotypes of HBV were determined directly by the size of PCR products.This nested PCR was used in the genotyping of HBVs in 15 patients with chronic hepatitis B.Results Of the 15 patients with chronic hepatitis B,7 (46.7%) were genotype B and 4 (26.7%) were genotype C,2 (13.3%) were genotype B+C.Conclusion With the application of this nested PCR,the predominant HBV genotypes in human are confirmed to be genotypes B and C.