实时荧光聚合酶链反应检测HBV基因型B～D方法的建立  

Establishment and application of real-time fluoriescene-based metry polymerase chain reaction for hepatitis B virus genotyping

作　　者：沈建坤[1] 周荣 王战会[1] 白培盛 马世武[1] 张克[1] 侯金林[1] 

机构地区：[1]南方医科大学南方医院感染内科,广东广州510515 [2]广州华银基因科技有限公司,广东广州510150

出　　处：《第一军医大学学报》2005年第6期630-632,共3页Journal of First Military Medical University

基　　金：国家973计划子课题(G1999054106)~~

摘　　要：目的建立实时荧光聚合酶链反应检测乙型肝炎病毒(HBV)基因型B^D方法并验证其准确性。方法比较GenBank中已发表的明确分型的143株HBV全序列,设计特异的组合引物探针,应用实时荧光聚合酶链反应HBV基因分型法对兰州地区128例慢性乙型肝炎患者血清标本进行基因型B、C、D检测,并随机对检测的基因型各3株标本进行S基因测序验证。结果128例标本中B型检出率为20.3%(26/128),C型71.9%(92/128),D型7.8%(10/128);18株HBV克隆标本S基因测序结果与本分型法完全一致。结论实时荧光聚合酶链反应HBV基因分型法能够简便、灵敏、快速、准确地鉴定HBV基因型,适宜用于HBV基因型的大规模临床和流行病学调查。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. Conclusion The real-time PCR method is convenient, highly sensitive, rapid and accurate, especially suitable in clinical and large-scale epidemiological studies.

关 键 词：肝炎病毒 乙型 基因型 实时荧光聚合酶链反应 

分 类 号：R446.5[医药卫生—诊断学]