基于数字PCR的转基因水稻LL62品系精准定量检测方法建立  

Establishment of Precisely Quantitative Method of Genetically Modified Rice LL62 Based on Digital PCR

作　　者：任怡菲[1,2] 高琴[2] 邓婷婷[3] 李想[2] 黄文胜[3] 陈舜胜[1] 陈颖[3] REN Yi-fei GAO Qin DENG Ting-ting LI Xiang HUANG Wen-sheng CHEN Shun-sheng CHEN Ying(School of Food Science, Shanghai Ocean University, Shanghai 201306 Shanghai Entry-Exit Inspection and Quarantine Bureau, Shanghai 200135 Chinese Academy of lnspection and Quarantine, Beijing 100176)

机构地区：[1]上海海洋大学食品学院,上海201306 [2]上海出入境检验检疫局,上海200135 [3]中国检验检疫科学研究院,北京100176

出　　处：《生物技术通报》2016年第8期69-76,共8页Biotechnology Bulletin

基　　金：科技部转基因生物新品种培育重大专项(2016ZX08012001-007)

摘　　要：为了促进转基因产品标识制度的顺利实施,对未获我国农业部批准的转基因水稻LL62品系进行精准定量检测方法研究,以保障我国进出口转基因农产品的检测监管。基于转基因水稻LL62品系外源插入片段和水稻基因组DNA的3'端旁侧序列设计数字PCR检测体系,建立了精准定量检测方法。结果显示,设计的引物探针反应效率高,基于此建立的数字PCR方法高度特异于转基因水稻LL62品系检测;方法可重复性好,生成微滴数相对标准偏差(RSD)值介于0.60%-11.11%;准确度高,3个混合样品(10%、1%和0.1%)测定值与真实值之间的偏差(bias)分别为0.12、0.09和0.10,相对标准偏差(RSD)介于0.09%-10.31%。建立的转基因水稻LL62品系微滴数字PCR定量检测方法简便快捷、特异性强、重复性好、准确度高,可准确有效地对农产品和食品中转基因水稻LL62成分进行精准定量分析。In order to promote the smooth implementation of labeling policy for genetically modified（GM）products,we study an precise quantitative detection method for the GM rice LL62 that has not been approved by the Ministry of Agriculture in China,aiming at guarantee the detection and supervision of GM agricultural products in export and import. A digital PCR（dPCR）detection system were designed based on the inserted exogenous fragment of LL62 and 3’flanking sequence of DNA of rice genome,and a precisely quantitative method was established. The results showed that the designed probes presented high efficiency,the developed dPCR method was highly specific for GM rice LL62 detection. The method was highly repeated,and the relative standard deviation（RSD）values of droplets’number ranged from 0.60%-11.11%. Results of the quantification of three blind samples showed that the bias between the true value and the measured value was 0.12,0.09 and 0.10,the RSD was 0.09%-10.31%,indicating that the accuracy was high. In conclusion,the established droplet dPCR method for GM rice LL62 detection is simple and convenient with high specificity,solid repeatability and high accuracy,and is suitable for precisely and effectively quantitative analysis of ingredients of GM rice or peeled rice LL62 in agricultural products and foods.

关 键 词：转基因水稻LL62品系 微滴数字PCR 品系特异性 定量 

分 类 号：S511[农业科学—作物学]