摘要
本研究根据赤羽病毒(AKAV)的S基因序列设计合成了引物,RT-PCR扩增得到目的基因片段并克隆到PGM-T载体得到质粒标准品。通过对荧光定量PCR反应条件的优化,建立了SYBR Green Ⅰ荧光定量PCR检测AKAV的方法。该方法的检测敏感性达到926拷贝/25μL,并具有较好的重复性和特异性。本试验方法的建立对于加强进出口牛AKAV的检验检疫具有十分重要的意义。
In the study, the primers were designed and synthesized according to the conservative S sequence of AKAV. The genes were amplified by reverse transcription polymerase chain reaction （RT-PCR） and cloned to the PGM-T vector. The reaction parameters were optimized to develop SYBR Green Ⅰ fluorescence quantitative PCR assay. It was shown that the fluorescence quantitative PCR assay could detect 926 copies/25μL of plasmid DNA and its specificity and reproducibility were very good. The method was very useful for laboratories working with early and rapid identification of AKAV for cattle importatim and exportation.
出处
《中国动物检疫》
CAS
2013年第8期70-74,共5页
China Animal Health Inspection
基金
福建省自然科学基金科技计划项目(2010IK016)
厦门市科技计划项目(3502Z2010012)
宁波市科技计划项目(2007C10057)
国家质检总局科技计划项目(2005IK049)
