电击法转化金针菇菌丝碎片表达报告基因eGFP及GUS的研究  

Electroporation of Mycelial Fragments to Express Reporter Genes eGFP and GUS in Flammulina velutipes

作　　者：刘建雨 张美彦[1] 张丹[1] 徐珍[1] 宋春艳[1] 李巧珍[1] 尚晓冬[1] 谭琦[1] 黄志勇 胡卫红 张引芳 赵静 王瑞娟[1] LIU Jianyu;ZHANG Meiyan;ZHANG Dan;XU Zhen;SONG Chunyan;LI Qiaozhen;SHANG Xiaodong;TAN Qi;HUANG Zhiyong;HU Weihong;ZHANG Yinfang;ZHAO Jing;WANG Ruijuan(Institute of Edible Fungi,Shanghai Academy of Agricultural Sciences,Key Laboratory of Edible Fungi Resources and Utilization（South）,Ministry of Agriculture,P.R.China,National Engineering Research Center of Edible Fungi,Shanghai Key Laboratory of Agricultural Genetics and Breeding,Shanghai 201403,China;Shanghai Xuerong Biotechnology Co.,Ltd.,Shanghai 201401,China;Shanghai Bright Esunyes Bio-tech Co.,Ltd,Shanghai 201400,China)

机构地区：[1]上海市农业科学院食用菌研究所农业部应用真菌资源与利用重点开放实验室国家食用菌工程技术研究中心上海市农业遗传育种重点实验室,上海201403 [2]上海雪榕生物科技股份有限公司,上海201401 [3]上海光明森源生物科技有限公司,上海201400

出　　处：《食用菌学报》2018年第2期23-28,共6页Acta Edulis Fungi

基　　金：上海市市级农口系统青年人才成长计划项目[沪农青字(2017)第1-13号];上海市食用菌产业技术体系建设专项资金[沪农科产字(2017)第9号]

摘　　要：以不完全酶解的金针菇(Flammulina velutipes)菌丝碎片作为转化受体,通过电击转化法将载体pYN6981导入金针菇单核体Dan3基因组中。结果显示:在复筛培养基上生长良好的转化子,其基因组中可扩增出潮霉素基因片段,通过激光共聚焦显微镜可观察到绿色荧光,GUS组织染色显示为蓝色,说明目的载体已经成功插入金针菇基因组中并获得表达。转化效率达16个转化子每10~6个菌丝碎片。有丝分裂稳定性分析显示80%的转化子可以稳定遗传。Using mycelial fragments of monokaric Flarnrnulina velutipes strain Dan3 from incomplete enzymatic digestion as the receptor, plasmid pYN6981 was transformed into the genome of F. velutipes via electroporation. The results showed that the transformed colonies grew well on the secondary screening plates, contained the hygromycin resistance gene fragment in their genomes, emitted green fluorescence under the confocal laser scanning microscopy, and displayed blue color under the GUS staining. These results suggested successful genomic insertion and expression of the plasmid. The transformation efficiency was 16 transformants per 106 mycelial fragments. The mitosis stability test showed that 80% of the transformants were genetically stable.

关 键 词：遗传转化 绿色荧光蛋白 GUS组织染色 

分 类 号：S646.15[农业科学—蔬菜学]