With conserved regions and regions of high variations, 16s rDNA is an important molecular basis for the biological species identification and system evolu- tion. The modem molecular biology with 16s rDNA as the primer can accurately re- veal the diversity of microorganisms species and inheritance, thereby 16s rDNA se- quence analysis has become the main basis for classification and identification of microorganisms. Having overcome the limitations of traditional microculture methods, this method is easy to operate, quick and accurate to detect with high sensitivity, making it widely apply to species identification, community comparative analysis, phytecoenogenesis and the assessment of population diversity. It is a objective classification method with high credibility.
[Objective] This study aimed to investigate the expression of exogenous gene in prokaryotes and eukaryotes. [Method] The expression vector pTYB2-WF harboring target gene was transformed into E. coli ER2566. IPTG was employed to induce the expression of phytase gene. The expression of phytase fusion protein was detected by SDS-PAGE, and the fusion protein was further purified. Phytase gene phyA was expressed in Pichia pastoris expression system. Yeast recombinant vector pPIC9K-phyA was constructed and transformed into P. pastoris GS115 to construct engineering strain GS115-pPIC9K-phyA. [Result] Phytase protein was ex-pressed under methanol induction. Enzyme activity assay indicated that the activity of phytase was 7.3 U/ml. P. pastoris engineering strain GS115-pPIC9K-phyA was successful y constructed. [Conclusion] Methanol yeast expression mechanisms play a certain role in molecular biology and industrial applications.
