摘要
WYN17132是我国自主研发的转am79-epsps基因耐草甘膦转基因玉米品种,已获得全国生产应用安全证书,但目前缺少相应的检测方法。为进一步落实我国转基因产品监测工作,本研究建立了针对该转化体特异性的实时荧光PCR检测方法。以转基因玉米WYN17132的外源插入序列与玉米基因组连接区域的侧翼序列为靶序列,设计多组特异性引物探针,筛选最佳引物探针组合,并系统评价该方法的特异性、检出限(LOD)和再现性。通过筛选,确定17132RB-qF3/qR3/qP3为最佳引物探针组合,该组合扩增片段为132 bp。本研究建立的WYN17132转化体特异性实时荧光PCR检测方法特异性良好,仅对目标转化体产生特异性扩增;方法灵敏度高,检出限可达0.05%;再现性稳定,适用性强。该检测方法能满足转基因玉米WYN17132的检测需求,为我国转基因玉米的监测监管和标识溯源提供了技术支撑。
WYN17132 is a glyphosate-tolerant transgenic maize variety with am79-epsps gene independently developed in China.It has obtained the national production and application safety certificate,but there is no corresponding detection method at present.In order to further implement the monitoring of genetically modified products in China,this study established a specific real-time PCR detection method for transgenic maize WYN17132 event.The insertion sequence of transgenic maize WYN17132 and the flanking sequence of the connected part of the maize genome were used as the target sequence to design multiple sets of specific primers and probes.The specificity,limit of detection(LOD),and reproducibility of the established method were systematically evaluated.After screening,17132RB-qF3/qR3/qP3 was selected as the optimal primer-probe combination,and the amplified fragment size was 132 bp.The developed specific real-time PCR detection method for WYN17132 demonstrated excellent specificity,producing amplification only for the target event.The method showed high sensitivity,achieving a LOD of 0.05%.It also exhibited stable reproducibility and strong applicability.The detection method could meet the detection requirements of transgenic maize WYN17132,and provide technical support for the monitoring,supervision and traceability of transgenic maize in China.
作者
咸若彤
缪青梅
彭城
陈笑芸
杨蕾
徐晓丽
魏巍
徐俊锋
李玥莹
汪小福
XIAN Ruotong;MIAO Qingmei;PENG Cheng;CHEN Xiaoyun;YANG Lei;XU Xiaoli;WEI Wei;XU Junfeng;LI Yueying;WANG Xiaofu(College of Life Sciences,Shenyang Normal University,Shenyang 110034,China;National Key Laboratory of Agricultural Product Quality and Safety,Key Laboratory of Traceability for Agricultural Genetically Modified Organisms,Ministry of Agriculture and Rural Affairs,Zhejiang Key Laboratory of Crop Germplasm Innovation and Utilization,Zhejiang Academy of Agricultural Sciences,Hangzhou 310021,China)
出处
《浙江农业学报》
北大核心
2025年第7期1397-1406,共10页
Acta Agriculturae Zhejiangensis
基金
浙江省自然科学基金重点项目(LZ23D030001)
农业生物育种重大项目(2022ZD0401908)。
