[Establishment and application of real-time fluoriescene-based metry polymerase chain reaction for hepatitis B virus genotyping].
OBJECTIVE
To explore a new method for determining hepatitis B virus (HBV) genotypes B-D with real-time fluorescence-based PCR.


METHODS
The PCR primers and probes were designed according to the analysis of 143 complete HBV genomes from GenBank and on the basis of a search for genotype-specific nucleotide sequences which were conserved in the 3 HBV genotypes. Real-time fluorescence-based PCR was performed for HBV genotyping of 128 samples collected from Lanzhou. Three samples of each genotype of B, C and D were randomly selected and their S gene was sequenced for confirmation of the results of PCR-based method.


RESULTS
Real-time PCR identified genotype B in 26 (20.3%), genotype C in 92 (71.9%), and genotype D in 10 (7.8%) cases. The sequencing results of the S gene of 18 PCR-produced clones were in complete consistence with those of fluorescence-based PCR.


CONCLUSIONS
The real-time PCR method is convenient, highly sensitive, rapid and accurate, especially suitable in clinical and large-scale epidemiological studies.
Shen
Jian-kun
JK
Department of Infectious Diseases, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China. shjk_2003@163.com
Zhou
Rong
R
Wang
Zhan-hui
ZH
Bai
Pei-sheng
PS
Ma
Shi-wu
SW
Zhang
Ke
K
Hou
Jin-lin
JL
D005470:Fluorometry / Q000379:methods; D016679:Genome, Viral / Q000235:genetics*; D005838:Genotype; D006515:Hepatitis B virus / Q000235:genetics*; D019694:Hepatitis B, Chronic / Q000821:virology*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods