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建立以Sanger测序为基础的H9N2亚型禽流感病毒全基因组测序方法。从GenBank和GISAID数据库中选取2000至2012年中国地区和中国国家流感中心病毒序列数据库中H9N2亚型禽流感病毒的8个片段基因序列,通过比对分析在相对保守的区域设计分段扩增引物,共16对。选用1株病例分离毒株和4株环境分离毒株对引物进行验证和进一步优化。优化后的16对引物能够有效扩增5株H9N2亚型禽流感病毒,仅个别反应出现非特异扩增。所有获得的目的片段仍能有效进行后续测序实验。建立了一种能够有效获得新型重配H9N2亚型禽流感病毒全基因组序列的Sanger测序方法,为该病原分子流行病学研究和开展风险评估提供技术支撑。
Abstract：To establish a simple method to amplify and sequencing the whole genome of avian influenza A(H9 N2)virus.The genes of avian influenza virus H9 N2 subtype in China from 2000 to 2012 were downloaded from database of NCBI(The National Center for Biotechnology Information)and GISAID(Global Initiative on Sharing All Influenza Data).16 primer pairs targeted to conserved regions of these gene sequences were designed.These primers were tested and optimized by 1 strain isolated from human respiratory tract and 4 strains isolated from environment.After optimization,the whole genome sequence of 5 avian influenza A(H9 N2)viruses were obtained with 16 primer pairs,although few primer pairs produced non-specific products,the target fragments could still be used for the following sequencing experiment efficiently.A Sanger protocol for generating the whole genome sequence of the reassortant avian influenza A(H9 N2)virus was established and technical support for molecular epidemiology and risk assessment of the virus was provided.
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基本信息:
DOI：10.13242/j.cnki.bingduxuebao.003303
中图分类号:R373.13
引用信息:
[1]何军,李晓丹,何兰,等.H9N2亚型禽流感病毒全基因组序列测定方法的建立[J].病毒学报,2018,34(01):40-44.DOI:10.13242/j.cnki.bingduxuebao.003303.
基金信息:
安徽省卫计委2017年科研计划项目(项目号:2017jk005),题目:安徽省H9N2亚型禽流感病毒的遗传进化研究~~
2018-01-24
2018-01-24
2018-01-24