Development of a TaqMan MGB RT-PCR for the rapid detection of H3 subtype avian influenza virus circulating in China

被引:7
|
作者
Teng, Qiaoyang [1 ,2 ]
Shen, Weixia [1 ]
Yan, Dawei [1 ]
Yan, Liping [1 ,2 ]
Li, Xuesong [1 ,2 ]
Li, Guoxin [1 ,2 ]
Yang, Jianmei [1 ,2 ]
Li, Zejun [1 ,2 ]
机构
[1] Innovat Team Pathogen Ecol Res Anim Influenza Vir, Shanghai 200241, Peoples R China
[2] Chinese Acad Agr Sci, Shanghai Vet Res Inst, Dept Avian Infect Dis, Shanghai 200241, Peoples R China
基金
国家高技术研究发展计划(863计划); 中国国家自然科学基金;
关键词
Minor groove binder; H3; subtype; Sensitivity; Detection; A VIRUSES; H9; VALIDATION; SWINE; ASSAY; PIGS; H5;
D O I
10.1016/j.jviromet.2015.02.025
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Previous studies demonstrated that the H3 avian influenza virus (AIV) in China is isolated most frequently from wild birds and live poultry markets. However, there is no subtype-specific real-time polymerase chain reaction (RT-PCR) available for the rapid and highly sensitive identification of H3 AIV. In this study, a TaqMan minor groove binder (MGB) probe and a pair of primers were designed based on a conserved region in the hemagglutinin gene of H3 AIV. These were used to generate an H3-MGB RT-PCR assay that recognizes only H3 AIV. The detection limit of the H3-MGB RT-PCR was 10 copies of DNA per reaction when 10-fold serial dilutions of T-H3HA plasmid were used as the template. This was 1000-times more sensitive than conventional RT-PCR. In experimental samples obtained from oropharyngeal swabs or cloacal swabs, the virus was detected in all ducks using H3-MGB RT-PCR, whereas only one duck tested positive for the virus in oropharyngeal swabs tested using conventional RT-PCR. The H3-MGB RT-PCR assay developed in this study is a sensitive and rapid tool for screening H3 AIV in China. (C) 2015 Elsevier B.V. All rights reserved.
引用
收藏
页码:64 / 69
页数:6
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