摘要
构建含潮霉素抗性基因和GFP基因的双元载体p CAMBgfp,以其为转化载体用农杆菌EHA105介导的方法对芒果胶孢炭疽菌Cg-8菌株进行遗传转化,以潮霉素抗性和GFP荧光来筛选阳性转化子。然后,通过离体接种芒果叶片和果实观察病斑大小筛选致病性缺陷转化子。结果获得500个阳性转化子,转化效率为平均106个孢子可获得400个左右同时具有潮霉素抗性和GFP荧光的阳性转化子,且其经多次在无潮霉素的培养基上传代后能得到稳定遗传。随机挑选其中13个突变体进行PCR检测,均可扩增出潮霉素基因目的条带,致病性分析从中筛选得到8个致病性减弱或缺失突变体。
The recombinant vector pCAMBgfp, containing hygromycin resistant gene and gfp reporter gene, was constructed and transformed into mango Colletotrichum gloeosporioides via Agrobacterium tumefaciens EHA105. Positive transformants were identified by both resistant to hygromycin and green fluorescence observation. Transformants defective in pathogenicity were screened out through inoculation assay on detached mango leave and fruit. Total 500 positive transformants were screened out from several transformation. The average transformation efficiency was up to 400 transformants per 106 conidia. All of the transformants tested remained stable in hygromycin resistant and gfp fluorescence after several generations of growth in the media absence of hygromycin. 13 candidate mutants were randomly selected for PCR detection of the gfp gene and 8 mutants shown pathogenicity reduction or lose.
出处
《热带农业科学》
2014年第8期47-51,共5页
Chinese Journal of Tropical Agriculture
基金
国家自然科学青年基金项目(No.31301824)
关键词
胶孢炭疽菌
遗传转化
突变体
致病性
Colletotrichum gloeosporioides
transformation
mutants
pathogenicity
