【作者】 山其木格；

【导师】 王炜； 崔春生；

【作者基本信息】 石河子大学 ， 生物化学与分子生物学， 2008， 硕士

【摘要】 β-葡聚糖酶的消化作用可促使饲料中β-葡聚糖的降解,从而在一定程度上消除其抗营养作用,提高饲料中营养成分的利用率。β-葡聚糖酶作为添加剂和对青贮饲料处理的研究及应用发展很快,范围也越来越广。经过大量实际应用,已普遍为饲料界所接受。目前在饲料工业中使用的酶制剂基本上是由大肠杆菌等微生物发酵产生的,发酵结束后需要将含酶发酵液经过一系列后处理操作,分离,加工成干物质才能应用于饲料生产中。这样工序繁杂,费时费力。若将β-葡聚糖酶在一个食品级宿主菌例如乳酸菌中进行表达,则不需进行酶制剂的发酵生产、分离和添加过程即可直接加入饲料中。本实验首先利用PCR方法从地衣芽孢杆菌中克隆到β-1,3-1,4-葡聚糖酶基因并测序,将该基因序列与其他地衣芽孢杆菌β-葡聚糖酶进行blast比对,分析其相互间的同源性。再将该基因与大肠杆菌表达载体pET32a进行连接,转化大肠杆菌βL21,得到了有效的表达。确定β-1,3-1,4-葡聚糖酶基因产物具有生物活性后,构建乳酸乳球菌表达载体pMG-glu,并电击转化乳酸乳球菌MG1363,最终得到有效表达的转化子。这为制备优良的青贮饲料专用乳酸菌剂奠定基础。更多还原

【Abstract】 β-glucanase can help degradeβ-glucan and effectively improve digestibility of animal diets. The research and application ofβ-glucanase as a silage additive are developing quickly, and the scope of utilization is on the rise. Through extensive practical application, it has been generally accepted by the animal feed additive industry. However, the production process of enzyme is quite complicated and the cost of it is high. Ifβ-glucanase could be expressed in GRAS host bacteria such as Lactis acid bacteria, it can be added directly to silage without any process such as production, fermentation and purification of the enzyme.In the research,β-1,3-1,4- glucanase from Bacillus licheniformis is cloned and sequenced, then Homology between the gene sequence and otherβ-1,3-1,4- glucanase from Bacillus licheniformis was analyzed. The gene ofβ-1,3-1,4- glucanase was ligated into pET32a and transformed in Escherichia coli BL21, eventually the effective expression was accomplished. After determined its effectiveness, the gene was used to construct a Lactococcus lactis expression vector pMG-glu, and transformed in Lactococcus lactis MG1363 by Electroporation, finally an effective transformant was obtained. This work has laid the foundation for preparing an excellent lactic acid bacteria addictive for silage.更多还原