脲基酰胺酶基因在黄酒酵母中的整合型表达

Constitutive expression of DUR1,2 gene in Chinese rice wine yeast

氨基甲酸乙酯是一种人类的潜在致癌物,在许多发酵酒中均有存在,主要来源于发酵过程中酵母代谢产生的尿素。以pYX212为载体,将脲基酰胺酶基因DUR1,2克隆到TPI强启动子和终止子之间的位点,再通过同源重组的方式将受强启动子调控的目的基因整合到黄酒酵母的基因组中,最终获得一株低产尿素的胞内脲基酰胺酶基因组成型高表达的黄酒酵母85#DUR1,2。在实验室规模的黄酒酿造实验中,85#DUR1,2产尿素量为8.34mg/L,比出发菌株降低了69.9%,贮存一段时间后的酒液中氨基甲酸乙酯含量比出发菌株降低了40.5%,而发酵性能、酒精度、总酸及氨基态氮与出发菌株无显著差异。

Ethyl carbamate（EC） is a potential carcinogen for human, which can be found in many fermented alcoholic beverages.The main precursor for EC is urea produced by yeast.The DUR1,2 gene which encoded urea amidolyase was cloned into plasmid pYX212 between the TPI promoter and terminator sites to form the DUR! ,2 expression cassette.The cassette was then integrated into the genome of the Chinese rice wine yeast 85#.Thus a urea-degrading Chinese rice wine yeast 85 was obtained.According to the laboratory scale rice wine brewing tests,urea was reduced to 8.34mg/L by yeast 85 69.9% less than the parental strain, while the ethyl carbamate level was 40.5% less than that of the parental strain after a period of storage.Meanwhile,85 was substantially equivalent to its parental strain in terms of fermentation ability, ethanol ,total acid and amino nitrogen.