YDL080C和LEU2基因敲除对工业黄酒酵母异戊醇生成量的影响  

Effect of YDL080C and LEU2 gene knockout on isoamyl alcohol production in industrial yellow rice wine yeast

作　　者：李童[1,2,3] 孙军勇[1,2,3] 吴殿辉[1,2,3] 李晓敏[1,2,3] 谢广发[3,4] 陆健[1,2,3,5] 

机构地区：[1]江南大学工业生物技术教育部重点实验室,江苏无锡214122 [2]江南大学粮食发酵工艺与技术国家工程实验室,江苏无锡214122 [3]江南大学生物工程学院,江苏无锡214122 [4]中国绍兴黄酒集团有限公司国家黄酒工程技术研究中心,浙江绍兴312000 [5]宿迁市江南大学产业技术研究院,江苏宿迁223800

出　　处：《食品工业科技》2015年第15期189-193,共5页Science and Technology of Food Industry

基　　金：食品加工过程安全控制的关键科学问题;973项目(2012CB720802);工业生物过程高效转化与系统集成的科学基础研究;973项目(2013CB733602);安全食品精深加工科技创新平台建设(2012B091400030);食品发酵过程酿酒酵母氮代谢物阻遏效应形成机制及其调控;国家自然科学基金重点项目(31130043);江苏高校优势学科建设工程资助项目;高等学校学科创新引智计划(111计划)资助项目(111-2-06);江南大学工业生物技术教育部重点实验室开放课题(KLIB-KF201308)

摘　　要：通过敲除生成途径中关键酶的编码基因,对异戊醇在工业黄酒酵母N85中的生成进行了研究。采用融合PCR技术,分别构建类丙酮酸脱羧酶基因(YDL080C)缺失组件"YDL080C-URA3-YDL080C"和β-异丙基苹果酸脱氢酶基因(LEU2)缺失组件"LEU2-URA3-LEU2",转化工业黄酒酵母N85尿嘧啶缺陷型单倍体Na-u(MATa ura3),获得单倍体工程菌Na-y和Na-l。将转化子与亲本分别进行实验室规模的黄酒发酵实验,结果 YDL080C缺失工程菌的异戊醇含量与亲本相比没有明显变化,说明类丙酮酸脱羧酶不是该途径关键酶;LEU2缺失工程菌的异戊醇含量降低16.16%,说明糖代谢途径存在,但只占小部分,而其他理化指标无明显差异。The production of isoamyl alcohol,was studied in industrial rice wine yeast N2,5 with the method of gene knockout.Disruption cassette of ＂ YDL080C- URA3- YDL080C ＂ and ＂ LEU2- URA3- LEU2 ＂ were constructed using fusion PCR, and then electro- transformed into the △ura3 haploid yeast strain （ MATa ura3 ）, respectively. Finally the engineered haploids Na- y and Na- l were obtained, Lab- scale fermentation of yellow rice wine was preceded with Na-y and Na-l, respectively.The results showed that the concentration of isoamyl alcohol was almost invariable between the engineered strain Na-y and the parental haploid. On the contrary, the content of isoamyl alcohol was reduced by 16.16% with the engineered strain Na-l.There was no difference in physical and chemical indexes between the engineered strain and parental strain.

关 键 词：异戊醇 工业黄酒酵母 类丙酮酸脱羧酶基因 β-异丙基苹果酸脱氢酶基因 基因敲除 

分 类 号：TS201.3[轻工技术与工程—食品科学]