基于重测序鉴定GbTMEM214基因在陆地棉基因组的插入位点  

dentification of GbTMEM214 insertion site in Gossypium hirsutum genome based on resequencing

作　　者：程俊凌 赵亮[2] 徐剑文[2] 刘剑光[2] 徐鹏[2] 徐珍珍[2] 郭琪[2] 王月平 赵君[2] 沈新莲[2] 陈全家[1] 肖松华[2] Cheng Junling;Zhao Liang;Xu Jianwen;Liu Jianguang;Xu Peng;Xu Zhenzhen;Guo Qi;Wang Yueping;Zhao Jun;Shen Xinlian;Chen Quanjia;Xiao Songhua(College of Agriculture,Xinjiang Agricultural University/Engineering Research Centre of Cotton,Ministry of Education,Urumqi 830052,China;Institute of Industrial Crops,Jiangsu Academy of Agricultural Sciences/Key Laboratory of Cotton and Rapeseed,Ministry of Agriculture and Rural Affairs,Nanjing 210014,China)

机构地区：[1]新疆农业大学农学院/棉花教育部工程研究中心,乌鲁木齐830052 [2]江苏省农业科学院经济作物研究所/农业农村部长江下游棉花与油菜重点实验室,南京210014

出　　处：《棉花学报》2024年第4期285-295,共11页Cotton Science

基　　金：农业生物育种重大项目(2023ZD04039-02-04);国家自然科学基金(31971904,31401727)。

摘　　要：【目的】利用农杆菌介导法获得了转GbTMEM214基因陆地棉品系,旨在明确转基因棉花株系中T-DNA插入位点的序列特征及检测方法,为其生物安全评价提供依据。【方法】基于基因组重测序技术,利用BLASTn将测序数据与陆地棉TM-1基因组序列进行比对分析,设计特异性引物通过聚合酶链式反应(polymerase chain reaction,PCR)验证插入位点。【结果】携带目标基因GbTMEM214的T-DNA整合在陆地棉基因组D13号染色体57019068~57019106 bp,T-DNA插入导致受体基因组37 bp的片段缺失;通过PCR扩增获得携带GbTMEM214基因的T-DNA插入位点的侧翼序列,由此建立了转GbTMEM214基因棉花的特异性检测方法。【结论】基于基因组重测序技术获得了转GbTMEM214基因棉花的T-DNA插入位点及侧翼序列信息,可为转基因棉花的生物安全评价提供技术参考。[Objective]GbTMEM214 transgenic Gossypium hirsutum line,obtained using Agrobacterium-mediated method,was used to clarify the sequence characteristics and detection methods of the T-DNA insertion site,and further promote its biosafety evaluation.[Methods]Based on the genome resequencing technology,the sequencing data was compared with the G.hirsutum standard line TM-1 genome sequence by BLASTn,and specific primers were designed to verify the insertion site by polymerase chain reaction(PCR).[Results]The T-DNA carrying the target gene GbTMEM214 was integrated into the position of 57019068-57019106 bp on chromosome D13 of G.hirsutum genome,resulting in 37 bp deletion of cotton genome.Combined with the flanking sequence of T-DNA insertion site obtained by PCR amplification,the specific detection method for GbTMEM214 transgenic cotton was established.[Conclusion]The T-DNA insertion site and flanking sequence of GbTMEM214 transgenic cotton was obtained based on genome resequencing technology,which can provide technical reference for biosafety evaluation of the transgenic cotton.

关 键 词：转基因棉花 GbTMEM214 重测序 插入位点 侧翼序列 

分 类 号：S562[农业科学—作物学]