Establish a new method of genotyping of hepatitis B virus by restriction pattern analysis of S ampicon

Objective A method was established for genotyping of hepatitis B virus (HBV),based on the restriction fragment length polymorphism(RFLP)created by BsrI, StyI,DpnI and HpaII action on an amplified segment of the S region. Methods 223 full-genomic sequences were analyzed and the aligned nucleotide and amino acid sequences of S gene, genotype specific regions were identified by the restriction enzymes, BsrI, StyI,DpnI and HpaII. Pre S PCR-RFLP genotyping method was applied to a number of serum samples from hepatitis B e antigen (HBeAg) positive and negative Chinese chronic HBV carriers. And in 90 samples the following genotypes were observed: 30B, 30 C, 30D. The method using S gene PCR-RFLP was confirmed to be correct by these 90 samples. Three samples of each genotype B, C and D were randomly selected and directly sequenced their S gene to confirm that HBV S gene PCR-RFLP genotyping method was correct disectly. Results The results of two PCR-RFLP HBV genotyping methods were coincide with that of S gene sequence. Conclusions The method for genotyping of hepatitis B virus (HBV), based on S gene RFLP is established to be highly sensitive, differential and accurate. The RFLP patterns are easy to be recognized because of its simplicity and singleness.