水稻OsNCED3基因的RNAi载体构建  

Construction of RNAi vector of OsNCED3 gene

作　　者：冯光秀[1] 陈惠[1] 

机构地区：[1]山西师范大学,生命科学学院植物生理实验室,山西临汾041000

出　　处：《生物学杂志》2012年第1期47-50,58,共5页Journal of Biology

基　　金：国家自然科学基金(No.30170236);重点实验室资助:中国农业大学生命科学研究中心植物生理与生物化学国家重点实验室

摘　　要：水稻OsNCED3基因是水稻抗逆过程中重要的基因之一。以水稻中花10号幼苗为材料,提取基因组DNA。设计引物扩增区段cDNA并引入相应的酶切位点,以基因组DNA作为模板,进行RNAi-OsNCED3顺式和反式目的片段的PCR扩增。将PCR产物连接到pMD19-T载体上,经酶切和PCR检测后进行测序。测序结果表明:RNAi-Os-NCED3顺式和反式目的片段均已正确的连接到pMD19-T载体上。然后将RNAi-OsNCED3顺式和反式目的片段通过酶切和连接,连接到含有发夹结构的质粒pFGC5941上。PCR及双酶切结果显示,构建的pFGC5941-OsNCED3即RNAi-OsNCED3载体结构完整。OsNCED3 gene is an important gene during stress tolerance. The genomic DNA was isolated from the seedings of Oryza sat# va ＂zhonghua NO. 10. ＂ The specific genomic DNA segment was chosen, the restriction site was inserted, the genomic DNA was used as a template for RNAi-OsNCED3 cis-and trans fragments of PCR amplification. The PCR amplification fragments was connected to the pMD19 - T vectors and sequenced after restriction enzyme digestion and PCR. The sequencing results showed that RNAi-OsNCED3 cisand trans fragments were correctly connected to the pMD19 - T vectors. The RNAi-OsNCED3 cis-and trans-fragment was digested by restriction enzyme and connected to the plasmid pFGC5941with a hairpin structure. PCR and double digestion verification showed that constructed pFGC5941-OsNCED3 vector （RNAi-OsNCED3） structural integrity.

关 键 词：水稻 RNAI OsNCED3 pFGC5941 

分 类 号：Q782[生物学—分子生物学] Q943.2