甜瓜响应甜瓜坏死斑点病毒侵染的cDNA文库构建  

Construction of Yeast Two-hybrid cDNA Library in Response to MNSV Infection in Melon

作　　者：吴会杰[1] 康保珊[1] 彭斌[1] 刘丽锋[1] 古勤生[1] WU Hui-jie;KANG Bao-shan;PENG Bin;LIU Li-feng;GU Qin-sheng(Zhengzhou Fruit Research Institute,Chinese Academy of Agricultural Sciences,Zhengzhou 450009,Henan,China)

机构地区：[1]中国农业科学院郑州果树研究所,河南郑州450009

出　　处：《中国蔬菜》2020年第9期36-40,共5页China Vegetables

基　　金：国家青年科学基金项目(31701941);河南省基础与前沿技术研究项目(152300410231,152300410232);中央级公益性科研院所基本科研业务费专项(1610192020501);国家西甜瓜产业技术体系病毒防控岗位项目(CARS-26-13)。

摘　　要：为解析甜瓜坏死斑点病毒(melon necrotic spot virus,MNSV)与寄主甜瓜的互作,筛选病毒编码的蛋白与寄主的互作因子,构建了MNSV侵染的甜瓜cDNA文库。在甜瓜品种白玫子叶期接种MNSV,待植株表现典型的系统坏死斑点症状后,收集症状明显的整株甜瓜幼苗,采用SMART(switching mechanism at 5′end of RNA transcript)技术合成cDNA,利用DSN(duplex-specific nuclease)对cDNA均一化处理后,首次构建了MNSV侵染甜瓜的cDNA文库并评价了文库的质量,文库容量为4.5×106 cfu,文库滴度为3.6×10^7 cfu·mL^-1,文库克隆重组率为100%,文库插入片段大小在500~2000 bp之间,说明文库质量很好。该文库的构建为解析MNSV与寄主互作及甜瓜响应MNSV的致病机制奠定了基础。In order to analyze the interaction between melon necrotic spot virus(MNSV)and host melon and screen the host factors interacted with the protein encoded by MNSV,this paper constructed cDNA library of melon infected with MNSV.During the cotyledon stage,melon variety‘Baimei’which is susceptible to MNSV was inoculated with MNSV.After the plant showed typical necrotic spot symptom,the melon seedlings with remarkable symptom were collected.The double strand cDNA was synthesized by switching mechanism at 5′end of RNA transcript(SMART)technique.Then the cDNA was treated by duplex-specific nuclease(DSN).For the first time,the paper constructed the melon yeast cDNA library and evaluated the quality of this library.The library capacity was 4.5×10^6 cfu.Its titer was 3.6×10^7 cfu·mL^-1.The library recombination rate was 100%.The size of inserting fragment of the library was 500-2000 bp,indicating that the library had very good quality.Construction of this library has laid a foundation for analyzing interaction between MNSV and host,and understanding melon response to MNSV pathogenic mechanism.

关 键 词：甜瓜 甜瓜坏死斑点病毒 CDNA文库 文库质量评价 

分 类 号：S436.5[农业科学—农业昆虫与害虫防治]