苦豆子凝集素基因植物表达载体的构建及其对烟草的转化

摘要

[Objective]To understand the role of Sophora alopecuroide lectin (SAL), SAL gene was used to construct the plant expressing vector and transformed tobacco for obtaining transgenic tobacco. [ Method ] Sophora alopecuroide lectin gene (SAL) was amplified from total RNA from Sophora alopecuroide leaf by RT -PCR technology. The amplified fragment was cloned into plant expression vector pCAMBIA1301 to generate recombinant plasmid pCAMBIA1301 - SAL. The vector was transformed into to tobacco by agrobacterium mediated method. Agrobacterium - mediated transient expression assay was evaluated. [ Result ] Plant expression vector pCAMBIA1301 -SAL was successfully constructed, and transformed into tobacco. Seventy - six resistance plants were obtained. PCR results showed that 27 plants were positive. [ Conclusion] Sophora alopecuroide lectin gene (SAL) was successfully expressed in tobacco. All of this laid foundations for the further study of plant disease resistance.%[目的]了解苦豆子凝集素基因(SAL)的功能,将该基因构建到植物表达载体并转化烟草,获得转基因植株.[方法]利用RT-PCR技术,提取苦豆子总RNA进行反转录得到cDNA,通过PCR扩增得到苦豆子凝集素基因SAL,并将其克隆到植物表达载体pCAMBIA1301上,产生重组质粒pCAMBIA1301-SAL.采用农杆菌介导法将重组质粒转化烟草,并进行分析鉴定.[结果]构建了植物表达载体pCAMBIA1301-SAL,转化烟草后经筛选获得76株转基因植株,经抗性筛选及PCR和RT-PCR鉴定,其中27株显示为阳性植株.[结论]苦豆子凝集素基因已经在烟草中成功表达,为进一步研究验证转基因烟草的抗病效果奠定了基础.