休哈塔假丝酵母木糖还原酶基因(xyl1)的克隆与序列分析  

Cloning and Sequence Analysis of the Xylose Reductase Gene(xyl1) from Candida shehatae

作　　者：杜仁鹏 黄守峰[1] 裴芳艺 葛菁萍[1] 平文祥[1,2] 

机构地区：[1]黑龙江大学生命科学学院/微生物省高校重点实验室,哈尔滨150080 [2]农业微生物技术教育工程研究中心,哈尔滨150500

出　　处：《中国农学通报》2015年第11期124-129,共6页Chinese Agricultural Science Bulletin

基　　金：国家自然科学基金面上项目"菌群演替与温水沤麻系统关键酶产生菌代谢组学特征的耦合机制"(31270534);国家自然科学基金面上项目"副干酪乳杆菌与枯草芽孢杆菌等菌群种间关系及协同合作策略的研究"(31470537);国家自然科学基金青年科学基金项目"酸菜发酵生态系统中细菌群落与代谢物组特征耦合机制"(31300355);黑龙江省高等学校科技创新团队"农业微生物发酵技术"(2012td009)

摘　　要：木糖还原酶氧化木糖生成木糖醇,处于木糖代谢的节点位置。酿酒酵母自身不具有木糖还原酶,不能利用木糖生产乙醇。因此,可以在酿酒酵母体内引入木糖还原酶基因xyl1,完善木糖代谢流,提高酿酒酵母的木糖利用率和乙醇得率。本研究利用PCR方法,根据木糖还原酶基因xyl1序列相似的特点,设计1对引物,以休哈塔假丝酵母基因组DNA和质粒PKT0150为模板,克隆得到了木糖还原酶基因xyl1,长度为1110 bp,有3个碱基缺失,ORF位于11-982位,全长为972 bp,编码323个氨基酸,缺失的3个碱基TTG位于ORF后10、11、12位。结果表明该片段为HDYXHT-20335木糖还原酶基因序列,为构建利用木糖高产乙醇的酵母菌奠定一定基础。Xylose can be oxidated to xylitol by xylosere duetase, which the reaction is the node position in thexylose metabolism. Xylose can＇t be utilized by Saccharomyces cerevisiae because of its lack of xylosereduetase. Therefore, the xyl1 should be inserted into the genome of Saccharomyces cerevisiae in order toconsummate xylose metabolic flux, and enhance xylose utilization rate as well as the ethanol yield by Saccharomyces cerevisiae. In this paper, according to the sequence similarity of xylose reductase, polymerasechain reaction（PCR） was used to amplify the xyl1 gene, a pair of primers were designed considering theCandida shehatae genome DNA and plasmid PKT0150 as template and finally 1110 bp cloning xyl1 of xylosereductase gene was obtained with three base missing, The ORF located in 11- 982, total length of 972 bp,encoding 323 amino acids after missing three bases TTG in the 10, 11, 12 of ORF. Results showed that thesegment was the xylose reductase gene sequencesas in HDYXHT- 20335 which laid the foundation for theconstruction of the yeast strain.

关 键 词：休哈塔假丝酵母 木糖还原酶 基因克隆 序列分析 

分 类 号：TS252[轻工技术与工程—农产品加工及贮藏工程]