Bt基因介导的大豆抗蚜虫表达载体的构建

Construction of Bt Gene-mediated Aphids-resistant Expression Vector in Soybean

[目的]构建用于大豆抗蚜虫研究的Bt基因介导的Cry1Ac植物表达载体。[方法]以含有Cry1Ac的质粒pRH201为模板进行PCR扩增,将PCR产物克隆至载体质粒pBS中,将重组质粒转化至大肠杆菌DH5α,并对重组质粒进行酶切和测序鉴定。经验证的Cry1A基因扩增产物双酶切后与pBI121载体连接转化,挑取重组子pBIC121进行BamHⅠ/SnaBⅠ双酶切鉴定。[结果]从pRH201质粒中扩增获得长约3.5 kb的Cry1Ac基因序列,对重组质粒pBSCRY的插入片段进行测定,确定了Cry1Ac片段的全长为3 534 bp,共编码1176个氨基酸。[结论]构建了Cry1A基因的植物表达载体,为大豆抗蚜虫转基因研究奠定了基础。

[ Objective ] The purpose was to construct the Bt gene-mediated plant expression vector Cry1Ac for research on the aphid-resistance in soybean. [ Method] With pRH201 plasmid containing Cry1Ac gene as the template PCR amplification was carried out. The PCR product was cloned to pBS piasmid and the recombinant plasmid was transformed into Escherlchia coll DH5α, which was identified by digestion and sequencing. After double-digested PCR products were connected and transformed with pBI121vector. The recombinant pBIC121 were identified with double enzyme digestion by BamH Ⅰ/SnaB Ⅰ. [ Result ] The Cry1Ac gene sequence about 3.5 kb was amplified from the plasmid PRH201. The insert fragments of recombinant plasmid PBSCRY were determined to make sure that the length of Cry1Ac was 3 534 bp, encoding 1 176 amino acids. [ Conclusion] In the experiment, expression vector of CrylA gene was constructed for soybean and laid the foundation for genetic research on aphid resistance in soybean.