作 者：
阮玲云;翁彩红;陈世友;陈文星;赖春芬;艾柳英;胡开辉;孙淑静
摘 要：
以p ET-28a-RFP质粒为模板,PCR扩增红色荧光蛋白(red fluorescent protein,RFP)基因,并克隆到真核表达载体p TE11上,置于RP27启动子调控之下,成功构建表达载体p TE11-RFP。采用PEG介导转化法,将p TE11-RFP转入银耳芽孢中。提取转化子基因组DNA,RFP基因特异性引物扩增获得与目的基因大小一致的特异条带;日光下肉眼观察转化子略显红色,荧光显微镜观察有明显的红色荧光。以上结果证明RFP基因已成功转入银耳芽孢并进行表达,RP27启动子可以调控外源基因RFP在银耳芽孢中的正确表达,为进一步研究外源基因在银耳芽孢生物反应器中的高效表达奠定了一定的基础。
译 名：
Construction of Plasmid Vector and Expression Identification of RFP Gene in Tremella fuciformis
作 者：
RUAN Lingyun;WENG Caihong;CHEN Shiyou;CHEN Wenxing;LAI Chunfen;AI Liuying;HU Kaihui;SUN Shujing;College of Life Sciences, Fujian Agriculture and Forestry University;
关键词：
Tremella fuciformis;;RFP gene;;Expression vector
摘 要：
The RFP(red fluorescent protein) gene was obtained by polymerase chain reaction(PCR) in which plasmid p ET-28a-RFP was used as a template, and cloned into p TE11 vector under the control of RP27 promoter. The recombinant plasmid p TE11-RFP was successfully constructed as confirmed by enzymatic digestion and DNA sequencing. The PEG mediated transformation method was performed to transfer plasmid DNA of p TE11-RFP into yeast-like conidia of Tremella fuciformis. The expected amplified bands appeared when the chromosomal DNA of the transformants was used as the templates for the PCR with the RFP-specific primers. The colonies showed pink in tube and distinct red fluorescence could be observed from the colonies of YLCs by fluorescence microscopy. These results indicated that RFP gene was integrated into the genome of T.fuciformis and was expressed successfully by the regulation of RP27 promoter which laid the foundation for foreign gene expression in YLCs of Tremella fuciformis.
