文档介绍：拟南芥 AtMYB61 基因的克隆及表达载体构建王云, 任永兵, 杨硕, 韩宇, 陶杨, 曹树青(合肥工业大学生物与食品工程学院,安徽合肥 230009) 摘要: 文章以拟南芥幼苗提取的总 RNA 为模板, 利用 RT-PCR 技术扩增获得 AtMYB61 基因的全长 cDNA 片段,再克隆到 pUCm-T 载体上,菌落 PCR 、酶切鉴定和 cDNA 测序结果表明成功克隆了拟南芥 AtMYB61 基因。从 AtMYB61 一 pUCm-T 载体上,用 BstEII 和 BglII 全双酶切切下目的基因片段, 将此基因片段连接到植物表达载体 pCAMBIA2301 中。菌落 PCR 和酶切鉴定结果表明成功构建了植物表达载体 pCAMBIA2301 一 AtMYB61 。利用电转化法将重组表达载体导人根癌农杆菌中,获得了携带 AtMYB61 基因的根癌农杆菌株,为转基因改良植物抗逆性和进一步研究 A ￡ MyB61 基因的抗逆分子机理奠定了基础。关键词:拟南芥; AtMYB61 基因;表达载体 Cloning of Arabidopsis thaliana AtMYB61 gene and the expression vector construction W ANG Yun , REN Yong-bing , YANG Shuo , HAN Yu, TAO Yang , CAO Shu-qing (School ofBiotechnology and Food Engineering ,Hefei University ofTechnology ,Hefei 230009 ,China) Abstract : Tota1 RNA was extracted from Arabidopsis thaliana seedlings and used as the template to amplify the full length of eDNA of AtMYB6 1 gene by RT_PCR technology , and the gene fragment was subsequently cloned into plant pUCm-T vector . The results of bacterial colony PCR , enzyme analysis and eDNA sequencing confirmed that the Arabidopsis thaliana AtM YB6 1 gene was essfully cloned . AtMYB61 gene was pletely from AtMYB61 一 pUCm-T vector by BstE 1]and Bgl Ⅱ, and the gene fragment was cloned into plant expression vector pCAMBIA2301 . The results of bacterial colony PCR and enzyme analysis showed the essfu1 construction of plant expression vector pCAM — BIA2301 一 AtM YB61 . In ad