利用流式细胞仪分选土壤中发生自然转化的阳性转化子  

Sorting positive transformants from natural transformation in soil with flow cytometer

作　　者：王玉萍[1] 黄巧云[1] 陈雯莉[1] 

机构地区：[1]华中农业大学农业微生物学国家重点实验室,武汉430070

出　　处：《华中农业大学学报》2016年第2期56-62,共7页Journal of Huazhong Agricultural University

基　　金：国家自然科学基金项目(41071180;41230854);国家"863"计划项目(2012AA101402)

摘　　要：为确定土壤中的DNA通过自然转化的方式发生水平基因转移的频率,将1个带有egfp和卡那霉素抗性基因Kanr的质粒DNA添加到自然土壤中,分离菌悬液,上流式细胞仪分选能发出绿色荧光且具有卡那霉素抗性的阳性转化子,确定土壤中发生自然转化的频率小于10^(-7)。分离的菌悬液通过培养基的富集,从39 736 426个细胞中分选获得了82个细胞,最终得到了纯化的阳性转化子,证实土壤中的DNA通过自然转化发生了水平基因转移。One plasmid DNA with egfp and Kanr was introduced into the natural soil. The positive transformants were isolated directly from the bacterial suspension with flow cytometer and sorted with green fluorescence and kanamycin resistance. The frequency of natural transformation was less than 10 7 in soil. After the enrich cultivation, 82 cells out of 39 736 426 were sorted. Positive transformants were finally obtained. The occurrence of horizontal gene transfer through natural transformation in soil was confirmed. Our studies break the lowest proportion limit of sorting with flow cytometry as well.

关 键 词：水平基因转移 自然转化 阳性转化子 流式细胞术 分选 

分 类 号：S151.9[农业科学—土壤学]