转基因马铃薯外源基因插入位点分析及检测方法的建立  

Analysis of Insertion Site of Transgenic Potato Exogenous Gene and the Establishment of Detection Method

作　　者：闫建俊 白云凤 左静静 李萌[2] 左敏 裴成成 Yan Jianjun;Bai Yunfeng;Zuo Jingjing;Li Meng;Zuo Min;Pei Chengcheng(College of Agriculture,Shanxi Agricultural University,Taiyuan,030031;Institute of Crop Germplasm Resources,Shanxi Academy of Agricultural Sciences,Taiyuan,03003)

机构地区：[1]山西农业大学农学院,太原030031 [2]山西省农业科学院农作物品种资源研究所,太原030031

出　　处：《分子植物育种》2020年第16期5361-5366,共6页Molecular Plant Breeding

基　　金：山西省青年科技研究基金项目(201701D221210);山西省农业科学院特色农业技术攻关项目(YGG17127);山西省农业科学院作物科学研究所青年基金(ZZQ1702)共同资助。

摘　　要：09-4为本实验室构建的抗马铃薯PRLV的RNAi基因和特异切割马铃薯PSTVd的核酶基因无标记双价表达载体p CAMBIA3301-DR-IS导入马铃薯中获得的转基因无性系。本研究通过染色体步移获取转基因无性系09-4的插入位点左边界旁侧序列,对获得的左边界序列进行分析,扩增获得片段大小为382 bp,包括转化载体序列119 bp及转基因马铃薯的旁侧序列263 bp;依据转化马铃薯的RNAi基因和左旁侧序列分别设计引物,扩增出大小为300 bp的片段,建立了09-4转基因无性系特异性标识和特异PCR检测方法,为转基因马铃薯的检测和身份识别提供技术基础。The transgenic potato line 09-4 is a genetically modified potato transformed with a marker-free binary vector harbouring an inverted repeat expression cassette of the IS and DR gene by Agrobacterium-mediated transfor-mation to the potato.Using genome walking PCR,left flanking sequence of the T-DNA integration site on potato genome was amplified.The flanking sequences to the left border was 382 bp in length,which contains 119 bp of T-DNA and 263 bp fragment of the potato genome.Based on the flanking sequences and the left boundary of the T-DNA sequences,the specific primes were designed respectively to amplify the fragment with a size of 300 bp.Using the primes,the event-specific PCR detection method for transgenic potato 09-4 lines established,the method developed in this study was suitable for event-specific detec-tion of the transgenic line 09-4.It provides technical basis for the detection and identification of transgenic potato.

关 键 词：转基因马铃薯 染色体步移 事件特异性检测 

分 类 号：S532[农业科学—作物学]