耐除草剂转基因大豆SHZD32-1数字PCR绝对定量方法的建立  

Development of Digital PCR Absolute Quantification Method for Herbicide-tolerant Genetically Modified Soybean SHZD32-1

作　　者：郑子繁 王颢潜 高佳奇 陈硕 柳方方 张秀杰 李亮[1,3] ZHENG Zi-fan;WANG Hao-qian;GAO Jia-qi;CHEN Shuo;LIU Fang-fang;ZHANG Xiu-jie;LI Liang(Biotechnology Research Institute,Chinese Academy of Agricultural Sciences,Beijing 100081,China;Development Center of Science and Technology,Ministry of Agriculture and Rural Affairs P.R.China,Beijing 100025,China;Institute of Quality Standard and Testing Technology for Agro-Products,Chinese Academy of Agricultural Sciences,Beijing 100081,China)

机构地区：[1]中国农业科学院生物技术研究所,北京100081 [2]农业农村部科技发展中心,北京100025 [3]中国农业科学院农业质量标准与检测技术研究所,北京100081

出　　处：《计量学报》2023年第3期415-421,共7页Acta Metrologica Sinica

基　　金：国家生物育种重大专项(2022ZD04020);中国农业科学院科技创新工程(CAAS-ASTIP-2023-IQSTAP)。

摘　　要：基于DNA单分子绝对定量的数字PCR技术,建立了该转化体的微滴式数字PCR(ddPCR)双重(转化体特异性基因和大豆内标准基因)定量方法,内容主要包括:优化双重ddPCR引物探针浓度和退火温度,考察特异性,确定线性动力学范围以及检出限(3 copies/μL)和定量限(15 copies/μL)等关键参数。转基因大豆SHZD32-1已获得生产应用安全证书“农基安正字(2019)”第293号,其ddPCR定量检测方法作为一种绝对定量的计量方法,简便高效、精准度高,将为标准物质定值、定量检测及规范生物安全领域数字PCR方法建立等提供技术支撑。Based on the digital PCR technology for absolute quantification of DNA single molecule,a droplet digital PCR(ddPCR)dual(transformant-specific gene and soybean internal standard gene)quantitative method for the transformant was established.The main contents include:optimize the double ddPCR primer-probe concentration and annealing temperature,examine the specificity,determine the linear kinetic range and critical parameters such as the limit of detection(3 copies/μL)and the limit of quantification(15 copies/μL).The genetically modified soybean SHZD32-1 has obtained the production and application safety certificate"Nongjianzhengzi(2019)"No.293.Its ddPCR quantitative detection method,as an absolute quantitative measurement method,is simple,efficient and accurate,and will provide technical support for the calibration of reference materials,quantitative detection and standardization of the establishment of digital PCR methods in the field of biosafety.

关 键 词：计量学 转基因检测 数字PCR 绝对定量方法 转基因大豆 标准物质 

分 类 号：TB99[一般工业技术—计量学]