植物乳杆菌α-葡萄糖基转移酶基因挖掘及表达研究

发布时间：2018-03-29 05:31

本文选题：乳酸菌　切入点：α-葡萄糖基转移酶　出处：《现代食品科技》2017年02期

【摘要】：α-葡萄糖基转移酶(α-GTF)是一类可以利用麦芽糖,异麦芽糖,O-甲基葡萄糖苷等为底物合成低聚糖的多功能性酶,因其功能多样性和独特性而备受关注。本文利用本团队测序的植物乳杆菌(FMNP01)全基因组信息,采用生物信息学方法分析获得α-葡萄糖基转移酶基因(α-gtf),同时根据NCBI中上传的乳酸菌全基因组信息中的α-GTF的DNA序列设计引物,利用PCR技术分别从植物乳杆菌、干酪乳杆菌和短乳杆菌中克隆得到3个α-gtf基因,其编码的氨基酸序列相似性分别为42%、29%和29%;将克隆得到的3个基因分别与表达载体pET-32a连接构建成表达质粒,转入大肠杆菌BL21表达宿主菌中进行诱导表达,产物经SDS-PAGE分析表明,在75 ku处都有一明显条带,与预期蛋白分子量一致,表明这3个基因成功获得了表达。同时对诱导剂浓度、诱导时间和温度进行优化,可望为以后相关研究奠定了基础。
[Abstract]:伪 -GTFs are a kind of multifunctional enzymes which can synthesize oligosaccharides from maltose, isomaltose O-methylglucoside and so on. The whole genome information of Lactobacillus plantarum sequenced by our team FMNP01 was used. The 伪 -glucosyltransferase gene (伪 -gtfFN) was obtained by bioinformatics, and primers were designed according to the 伪 -GTF DNA sequence in the whole genome information of lactic acid bacteria uploaded from NCBI. The primers were obtained from Lactobacillus plantarum by PCR technique. Three 伪 -gtf genes were cloned from Lactobacillus casei and Lactobacillus brevis, with the amino acid sequence similarity of 422.2% and 29.9%, respectively. The product was induced to express in Escherichia coli BL21 expression host strain. SDS-PAGE analysis showed that there was an obvious band at 75 ku, which was consistent with the expected protein molecular weight, indicating that the three genes were successfully expressed. The optimization of induction time and temperature is expected to lay a foundation for further research.
【作者单位】：华南农业大学食品学院食品生物技术研究所;广东省微生态制剂工程技术研究中心;
【基金】：国家自然科学基金资助项目(31272217,31372116) 广东省科技计划项目(2015030313425,2015A030310225,2015A020209121)
【分类号】：Q55;Q78

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