产黄青霉简单高效遗传转化法——电激转化法

A simple and high-efficient electroporation method for Penicillium chrysogenum

目的研究产黄青霉(P.chrysogenum)电激转化方法,建立产黄青霉快速、高效的遗传转化方法。方法应用美国Bio-Rad电转仪,将外源DNA转化入产黄青霉菌丝中,研究了菌丝培养时间、电场强度、DNA类型等对遗传转化的影响。结果菌丝培养24h,处于旺盛生长阶段,电场强度为6000V/cm,电阻200Ω,电容20μF,环状质粒DNA较线状能获得转化效率稍高,1μg DNA获得的克隆数可达500个。结论应用电激转化方法对产黄青霉进行遗传转化属首次报道,特别是可直接将目的片段与T-DNA相融合,转化入产黄青霉中,该方法操作简单,快速高效。

Objective To study a simple and high-efficient electroporation method for P chrysogenum. Methods The exogenous DNA was electroporated into the mycelium ofP chrysogenum with Bio-Rad Gene Pulser. The cultural time for P chrysogenum mycelium, electric field intensity and DNA type were studied. Results The higher transformation efficiency （500clones/μg DNA） was obtained by electroporated plasmid DNA into mycelium cultivated for 24h at the condition of 20μF, 200Ω 6000V/cm. Conclusion The electroporation method for P chrysogenum was reported firstly. The interested gene fused with T-DNA from Angrobacteria cab be electroporated into P chrysogenum directly. The method is simple and hi^h-efficient.