产紫杉醇真菌N8菌株URA-3基因的敲除  

Disruption of URA-3 gene of a paclitaxel-producing fungus N8

作　　者：颜菲[1] 

机构地区：[1]天津医科大学生物医学工程学院,300070

出　　处：《国际生物医学工程杂志》2014年第5期257-261,F0003,共6页International Journal of Biomedical Engineering

基　　金：国家自然科学基金资助项目（30970084）

摘　　要：目的 为了解决产紫杉醇小孢拟盘多毛孢真菌N8基因操作的筛选标记缺乏的问题,构建N8菌株营养缺陷型菌株.方法 通过基因同源重组的方法定向敲除N8菌株中尿嘧啶合成途径中关键基因URA-3基因,然后利用分子生物学方法和添加一定浓度5-氟乳清酸（5-FOA）、尿嘧啶的基本培养基筛选获得转化子.结果 尿嘧啶营养缺陷型菌株在含有5-FOA和尿嘧啶的培养基上可以正常生长而野生型N8菌株无法生长.结论 成功构建产紫杉醇真菌N8菌株的尿嘧啶营养缺陷型菌株,可为其后续的基因功能研究奠定基础.Objective In order to get genetic markers,an auxotrophic paclitaxel-producing fungus named Pestalotiopsis malicola N8 strain was isolated by genetic modification.Methods Based on the homologous recombination,URA-3 which is the key gene for uracil synthetic route of Pestalotiopsis malicola N8 strain was knocked out.The transformants were screened by minimal medium with the combination of 5-fluoroorotic acid （5-FOA） and uracil.Results The results showed that the uracil auxotrophic strain was able to grow in the minimal medium containing 5-FOA and uracil while the wild type strain was not.Conclusions The uracil auxotrophic strain can be used as a new selection marker for future gene function studies of N8 strain.

关 键 词：产紫杉醇真菌 URA-3 基因敲除 

分 类 号：Q78[生物学—分子生物学]