Construction of Plasmid Vector and Expression Identification of RFP Gene in Tremella fuciformis
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.