Copy number and zygosity determination of transgenic rapeseed by droplet digital PCR  

作　　者：Yuhua Wu Jun Li Xiaying Li Jingang Liang Yunjing Li Xinhua Zeng Gang Wu 

机构地区：[1]Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Oil Crops Research Institute,Chinese Academy of Agricultural Sciences, Wuhan, China [2]Science and Technoloty Center of Ministry of Agriculture, China

出　　处：《Oil Crop Science》2017年第2期84-94,共11页中国油料作物学报(英文版）

摘　　要：Establishing an accurate and rapid method for copy number and zygosity determination can accelerate genetic engineering research process. In this study, droplet digital PCR (ddPCR), an emerging DNA absolute quantification technology, was used to identify single-copy homozygous transgenic lines from a batch of T0 transgenic rapeseed harboring 11 exogenous elements. Copy number of exogenous gene was evaluated in T0 generation based on calculated ratio between transgene and reference CruA gene, single-copy transformants were selected for selfing followed by subsequent zygosity analysis. Single-copy homozygous transgenic plants were successfully screened out in T1 generation by ddPCR.Segregation analysis with T2 seedlings verified that identification results of ddPCR were accurate and reliable. This study provides a novel rapid and accurate method for copy number and zygosity determination in transgenic rapeseed which overcomes disadvantages of traditional Southern analysis and recently developed real-time quantitative method.

关 键 词：DROPLET DIGITAL PCR (ddPCR) copy number ZYGOSITY homozygotes transgenic RAPESEED 

分 类 号：S565.4[农业科学—作物学]