鲁氏酵母3-磷酸甘油脱氢酶基因(ZrGPD1)在粉状毕赤酵母中的表达

Heterologous Expression of Zygosacharomyces rouxii ZrGpd 1 in Pichia farinosa

为探索在野生型粉状毕赤酵母(Pichiafarinosa)中整合表达来源于耐高渗鲁氏酵母(Zygosacharomycesrouxii)的3-磷酸甘油脱氢酶基因(ZrGPD1)以提高产甘油能力的可行性,应用PCR方法从P.farinosa的染色体中扩增出乳清苷酸脱羧酶基因(URA3)片段,以此作为同源整合的靶序列,构建了整合型表达载体pUR-ZG。电击转化粉状毕赤酵母,以抗生素Zeocin为筛选标记,获得转化子pfa-gu,摇瓶发酵结果表明以P.farinosa作为对照菌株,发酵72h后,转化子pfa-gu的生物量和甘油含量均高于对照菌株,其中甘油含量达到37g/L,比对照提高了30%。因此,在P.farinosa中异源表达ZrGPD1能够提高细胞的产甘油能力和对渗透压的调节能力。

To examine the effects of heterologous expression of ZrGPD1 （encoding glycerol 3-phosphate dehydrogenase ） cloned from osmotolerant yeast Zygosacharomyces rouxii on glycerol production in wild Pichia farinosa, the URA3 gene was amplified from P. farinosa as the homology integrative region. A recombinant plasmid （pUR-ZG） was constructed then transformed into P. farinosa by electroporation. The transformant pfa-gu was obtained by the selectable marker ZeocinTM. Primary results showed that the biomass of pfa-gu was higher than the wild type in the flask and after 72h fermentation the concentration of glycerol of pfa-gu was 37g/L enhanced 30% in comparison with the wild type. It is concluded that heterologous expression of ZrGPD1 is useful for increasing glycerol production and the ability of osmoregulation in P. farinosa.