产黄青霉工业生产菌种基因报告系统的构建及启动子效率的评价

本研究以四种不同物种来源,不同代谢途径功能基因的启动子为材料,分别构建了以产黄青霉异青霉素N合成酶(IPNS)基因pcbC的启动子Pipns、构巢曲霉3-磷酸甘油醛脱氢酶基因gpdA的启动子PgpdA、构巢曲霉色氨酸合成基因trpC的启动子PtrpC、粗糙脉胞霉氨基酸合成交叉途径控制基因cpc的启动子Pcpc为启动子,腐草霉素(phleomycin)抗性基因为报告基因,构巢曲霉色氨酸合成基因trpC的终止子TtrpC为终止信号的丝状真菌转基因质粒,建立了产黄青霉工业生产菌种启动子筛选、评价体系,调查了这四种启动子的强弱。结果表明选择性标记基因受强启动子驱动可以提高产黄青霉工业生产菌种的转基因效率。研究也显示这四种启动子的强弱的顺序为PgpdA、Pcpc、Pipns和PtrpC。

CONSTRUCTION OF GENE REPORT SYSTEM FOR PENICILLIUM CHRYSOGENUM INDUSTRIAL STRAIN AND EVALUATION OF PROMOTER ACTIVITY

The promoter screening and evaluation system was constructed with phleomycin resistance gene (Sh ble) as the dominant selection marker and the reporter gene and the terminator of the trpC gene as the terminator of the reporter gene. The promoter activity of PgpdA, PtrpC, Pcpc-1, and Pipns, which came from the glyceraldehyde-3-phosphate gene of Aspergillus nidulans, the tryptophan biosynthetic pathway of Aspergillus nidulans, the general regulatory gene of cross-pathway control for amino acid metabolite in Neurospora crassa, and isopenicillin N synthetase gene of Penicillium chrysogenum, respectively, were evaluated with this system. The results showed that the transformation efficiency of the industrial strain Penicillium chrysogenum can be improved while dominant selection marker gene was driven by high efficiency promoter. The results also showed that the strength order of these four promoters was PgpdA, Pcpc, Pipns, PtrpC.