引用本文:[点击复制]
[点击复制]
【打印本页】【下载PDF全文】 查看/发表评论下载PDF阅读器关闭

←前一篇|后一篇→

过刊浏览    高级检索

本文已被:浏览次   下载 本文二维码信息
码上扫一扫!
兼抗玉米矮花叶病毒及粗缩病毒无选择标记 siRNA复合表达载体的构建
张姣,赵阳,甘德芳
0
(安徽农业大学园艺学院,合肥 230036;安徽农业大学生命科学学院,合肥 230036)
摘要:
根据GenBank和MaizeSeq等数据库中的玉米矮花叶病毒和粗缩病毒外壳蛋白基因(CP)全序列设计特异性引物,RT-PCR扩增玉米矮花叶病毒及粗缩病毒的CP基因特异性RNA干扰片段,分别构建反向重复片段并串联成pBluscript + SM;Hind III-EcoR I 双酶切2T-DNA真核表达载体pDTB,插入Ubiquitin启动子及Nos 终止子,构建pDTBU;串联的反向重复片段插入pDTBU,构建兼抗玉米矮花叶病毒和粗缩病毒的siRNA复合表达载体pDTBUSM。酶切结果显示,目的片段均正确插入到相应的载体中。本研究对开展抗病毒RNA干扰调控技术研究,创建高抗病毒玉米新种质具有重要的理论和实践意义。
关键词:  玉米矮花叶病毒  玉米粗缩病毒  外壳蛋白  载体构建  siRNA
DOI:CNKI: 34-1162/S.20110303.0953.016
基金项目:国家高技术研究与发展计划(863)项目(2008AA10Z408)和安徽省高校省级科研项目(KJ2011A110)共同资助。
Construction of marker-free siRNA complex expression vector against MDMV and MRDV
ZHANG Jiao,ZHAO Yang,GAN De-fang
(School?of?Horticulture, Anhui Agricultural University, Hefei 230036;School?of?Life?Science, Anhui Agricultural University, Hefei 230036)
Abstract:
Based on the coat protein gene sequence of MDMV and MRDV in GenBank and MaizeSeq database, six pairs of specific primers were designed and six specific fragments were amplified by RT-PCR to prepare siRNA that corresponded to part of the MDMV CP or MRDV CP gene. In the first cloning step, an inverted repeat sequence of pUCCRNAi + 2 F was constructed. Next, two inverted repeat sequences were inserted into pBluscript SK in series to generate pBluscript + SM. Meanwhile, Ubiquitin promoter and Nos termination were cloned into pDTB to generate pDTBU. In the third step, pDTBU and pBluscript + SM plasmids were digested and joined to generate pDTBUSM. The study presented here provides a valuable tool for plant viral control using RNAi and the PTGS approach.
Key words:  maize dwarf mosaic virus (MDMV)  maize rough dwarf virus (MRDV)  coat protein (CP)  vector construction  siRNA

用微信扫一扫

用微信扫一扫