The basic parts of our design include the PRM1 promoter, the MIT1 promoter, the AOX1 promoter, the PRM1 gene (transcription factor for PAOX1, PPRM1 and PMIT1), the MIT1 gene (transcription factor for PAOX1 and PPRM1), and the yEGFP3 gene. All except for yEGFP3 are homogenous to the P. pastoris GS115 strain. Composite parts are constructed from these parts and are respectively PAOX1-GFP, PMIT1-PRM1, PPRM1-MIT1 and PMIT1-PRM1-PPRM1-MIT1. All composite parts are to be reintegrated into wild type P. pastoris GS115 (for PMIT1-PRM1-PPRM1-MIT1, it is separate integration of PMIT1-PRM1 and PPRM1-MIT1). In the strain containing only PAOX1-GFP, yEGFP3 is the reporter gene that reflects AOX1 activity. The rest three composite parts shall each be integrated into the PAOX1-GFP strain, so that the up regulation of PAOX1 can be compared with the wlid type. Up-regulation is achieved via the recombination of the homogenous transcription factors and promoters so that the transcription factors are dynamically over-expressed.
