Construction of yeast one-hybrid library and screening of transcription factors regulating FPS expression in Ganoderma lucidum / 中草药

ABSTRACT

Objective: To screen the upstream regulatory transcription factors of farnesyl diphosphate synthase (FPS) in the triterpenoid synthesis pathway in Ganoderma lucidum. Methods: In this study, the FPS promoter was cloned and connected to the pAbAi plasmid to construct bait vector pAbAi-FPS, which was transformed into Y1H yeast competent cells to construct bait yeast. The yeast one-hybrid cDNA library was constructed by using SMART technology, then the purified ds-cDNA and pGADT7-Rec were co-transformed into bait yeast strain to screen the upstream transcriptional regulatory factors of PFS. Results: The bait vector containing pAbAi-FPS was constructed and the bait strain was screened, the cDNA library was constructed and transformed to the bait strain. A total of 37 positive clones were screened and sequenced. The sequences of conserved domain were predicted and performed blast search against the whole-genome database to identify their function. As a result, a total of 18 upstream regulatory factors were screened out including three transcription factors, five ribosomal proteins, and 10 other transcription regulators. Conclusion: The results indicated that transcription factors GlSNF2, GlMHR, and GlZn2Cys6 were candidate genes for regulating the expression of FPS, and this study offered data for further study on the regulation mechanism of FPS expression.