摘要
为建立牛源乙肝病毒(HBV)套式PCR检测方法,本研究根据GenBank中人HBVS基因,设计2对引物,通过对PCR反应条件进行优化,建立检测牛源HBV的套式PCR检测方法,并进行S基因片段序列同源性和系统发育进化树分析。结果表明:该方法第1次PCR扩增的敏感性是1.00pg,第2次PCR敏感性是0.32fg,扩增产物分别为509bp、240bp,而牛病毒性腹泻病毒、丝状支原体丝状亚种SC型、牛结核分枝杆菌的扩增结果均为阴性。牛源HBVS基因片段序列与GenBank中人HBVS基因进行比对,核苷酸同源性98.8%～99.4%,氨基酸序列同源性97.1%～98.3%。研究表明建立的牛源HBV套式PCR具有很好的特异性和敏感性,可用于牛源HBV的检测,并对进一步研究牛源HBV与人HBV之间的关系奠定了基础。
A nested PCR assay for detection of hepatitis b-type virus（HBV） in cow was established using specific primers derived from human HBV sequence.The assay could amplify two specific bands of 509 bp and 240 bp from HBV DNA at a sensitivity of 1.00 pg in the first amplication and 0.32 fg in the second amplification.None of PCR products were amplified from BVDV,MmmSC and Mycobacterium bovis.Sequence comparison and phylogenetic tree analysis showed that the S gene sequences of HBV of milk cow and human HBV shared 98.8%-99.4% identities at nucleotide level,and 97.1%-98.3% identities at amino acid level.The nested PCR assay was specific and sensitive,which could be an effective tool for detection of HBV of milk cow.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2010年第7期542-545,共4页
Chinese Journal of Preventive Veterinary Medicine
基金
贵州省优秀教育人才省长专项资金项目(黔省专合子[2005]255号)
贵州省重大专项(黔科合重大专项字[2006]6002)
关键词
牛源HBV
人HBVS基因
同源性分析
套式PCR
hepatitis b-type virus of milk cow
human hepatitis b-type virus
homology analysis
nested PCR
