转基因玉米Bt11品系特异性荧光RPA检测  

Specific Detection of Genetically Modified Maize Bt11 with Novel Realtime RPA Assay

作　　者：李凯[1] 金芜军[1,2] 李亮 苗朝华[1,2] 刘卫晓 宛煜嵩[1,2] 

机构地区：[1]中国农业科学院生物技术研究所,北京100081 [2]农业部转基因植物用微生物环境安全监督检测中心(北京),北京100081

出　　处：《分子植物育种》2017年第11期4741-4745,共5页Molecular Plant Breeding

基　　金：转基因生物新品种培育重大专项项目(2013ZX08012-001;2018ZX08012-001)资助

摘　　要：重组酶聚合酶介导的等温扩增(recombinase polymerase amplification,RPA)作为一种新型等温扩增技术,具有反应时间短、灵敏度高、特异性强等特点,在转基因检测领域得到了广泛关注。本研究针对转基因玉米Bt11外源插入链接区域设计筛选了特异性引物和探针,建立了快速准确的荧光RPA检测方法,在39℃条件下,20 min内完成扩增并且能够实时监测,可以准确有效地检测到50个拷贝的靶标片段。而且,对于背景复杂的植物基因组DNA能够得到准确的扩增曲线。本研究不仅可以满足常规实验室检测,通过RPA便携式仪器,还可以满足转基因植物现场检测,具有广阔的应用前景。Recombinase polymerase amplification（RPA） is a novel isothermal amplification technology in genetic modified organism（GMO） detection area, and receive great attention. The reaction finished within short time and keep high sensitivity and specificity. In this paper, based on the genetically modified（GM） maize Bt11, we designed specific RPA primers and established real-time RPA method for screening and detecting. This method amplified the target DNA in vitro within 20 minutes under the temperature of 37℃ to 42℃ and reliably detection as few as 50 copies of the GM maize Bt11. In addition, we can receive accurate results under complex plant genome.Equipped with portable testing instruments, detection of GM maize can be done in fields, which has broad application prospects.

关 键 词：等温扩增 实时荧光RPA 转基因检测 Bt11 

分 类 号：S513[农业科学—作物学]