动物医学进展Issue(11)：1-6,6.
水貂 IFN-α、IFN-β及 IFN-γmRNA 荧光定量RT-PCR 检测方法的建立及应用
Establishment and Application of Real-time RT-PCR for Detection of Mink Interferon-α,Interferon-βand Interferon-γmRNA
摘要
Abstract
In this study,the methods of SYBR GreenⅠreal-time quantitative RT-PCR assay for detection of mink IFN-α,IFN-βand IFN-γ(MiIFN-α,MiIFN-βand MiIFN-γ)mRNA were established using four pairs of specific primers designed according to the MiIFN-α,MiIFN-βand MiIFN-γgene sequences,with the mink glyceraldehyde-3-phosphate dehydrogenase (MiGAPDH)mRNA as an internal control.The target genes were amplified by RT-PCR and inserted to pMD18-T vector as the standard recombinant plasmids for the establishing the real-time PCR standard curves.The results showed that the Ct values of MiIFN-α,MiIFN-β,MiIFN-γand MiGAPDH mRNA had good linear relationships (R2 =1.00)with the standards from 10 copies/μL to 107 copies/μL,and the melting curve showed a single peak.And the sensitivity assay results showed that the genetic test minimum value ware 10 copies/μL.The coefficient variations of inter-assay and intra-assay were both less than 4.5%.The result of clini-cal test showed that the relative expressions of MiIFN-α,MiIFN-βand MiIFN-γreached the highest level.The ex-pressions of MiIFN-αwas 100 times higher than that of MiIFN-βand MiIFN-γ and increased significantly during the infection period.The detection methods provide an effective tool for quantitative analysis of mink IFN-α,IFN-βand IFN-γmRNA.关键词
SYBR GreenⅠ/实时荧光定量PCR/水貂/α干扰素/β干扰素/γ干扰素Key words
SYBR GreenⅠ/real-time PCR/mink/IFN-α/IFN-β/IFN-γ分类
农业科技引用本文复制引用
王洋,张蕾,薛向红,史宁,白雪,徐淑娟,范思宁,凌明玉,李欣彤,闫喜军,胡博,张海玲,鲁荣光,吕爽,刘昊,孙彦刚,马凡舒,赵建军..水貂 IFN-α、IFN-β及 IFN-γmRNA 荧光定量RT-PCR 检测方法的建立及应用[J].动物医学进展,2015,(11):1-6,6.基金项目
吉林省科技发展计划项目（20130206026NY，20140520172JH）；吉林市杰出青年项目（2013625018）；吉林市科技发展计划 （）
