转g10-epsps基因耐除草剂大豆ZUTS-33转化体特异性检测方法的建立  

Establishment of Transformant-specific Detection Method for the Herbicide-tolerant Transgenic Soybean Event ZUTS-33 Harboring the g 10-epsps Transgene

作　　者：缪青梅[1] 赵杨 徐晓丽[1] 徐俊锋[1] 汪小福[1] 陈笑芸[1] MIAO Qingmei;ZHAO Yang;XU Xiaoli;XU Junfeng;WANG Xiaofu;CHEN Xiaoyun(Institute of Agro-product Safety and Nutrition,Zhejiang Academy of Agricultural Sciences,Hangzhou 310021,China;College of Life Sciences,Zhejiang University,Hangzhou 310058,China)

机构地区：[1]浙江省农业科学院农产品质量安全与营养研究所,杭州310021 [2]浙江大学生命科学学院,杭州310058

出　　处：《生物技术进展》2020年第6期696-703,共8页Current Biotechnology

基　　金：浙江省自然科学基金项目(LGN18C200029)。

摘　　要：转g 10-epsps基因耐除草剂大豆ZUTS-33是由浙江大学研发的耐除草剂大豆品系,目前已进入生产性试验阶段。到目前为止尚无文献报道对该转基因新品种的检测方法,因此亟需建立精准的定量检测方法为农业转基因生物安全管理提供技术支持。根据耐除草剂大豆ZUTS-33品系外源基因插入位点特异序列设计引物和TaqMan探针,利用优化的实时荧光定量PCR检测方法评价该引物对和探针的特异性、准确度、精确度和重复性,并确定此检测方法的检测极限(limit of detection,LOD)和定量极限(limit of quantity,LOQ)。实验结果显示,研究所建立的转基因大豆ZUTS-33转化体特异性实时荧光定量PCR检测方法具有高度的品系鉴定特异性,准确度、精确度均符合要求,重复性较好,且检测方法的LOD达到20拷贝,LOQ达到40拷贝。研究结果为转g 10-epsps基因耐除草剂大豆ZUTS-33的身份识别和检测监测提供了有效的方法。ZUTS-33 is the transgenic soybean line developed by Zhejiang University with the modified agronomic trait of herbicide tolerance,which has entered the stage of safety assessment before commercial release.So far,there is no literature report on the detection method of this new transgenic variety,so it is urgent to establish an accurate quantitative detection method to provide technical support for the safety management of agricultural genetically modified organisms.Primers and TaqMan probes were designed according to the specific sequences of the transgene and the insertion site of the T-DNA in herbicide-resistant soybean ZUTS-33 genome.The specificity,accuracy,precision and repeatability of the primer pairs and probes were evaluated by the optimized real-time fluorescence quantitative PCR detection method,and the detection limit of detection(LOD)and quantitative limit(LOQ)of this detection method were determined.The experimental results showed that the established real-time fluorescence quantitative PCR detection method for transformant specificity of transgenic soybean ZUTS-33 had high specificity of line identification,and accuracy and precision met the requirements,and repeatability was good.LOD and LOQ of the detection method reached 20 copies and 40 copies respectively.The results provided an effective method for identification,detection and monitoring of transgenic soybean ZUTS-33 with the g 10-epsps gene.

关 键 词：转基因大豆 g 10-epsps基因 ZUTS-33 实时荧光定量PCR 定量检测 

分 类 号：S565.1[农业科学—作物学]