贝莱斯芽孢杆菌Bacillus velezensis YB15β-葡聚糖酶的抑菌作用与基因克隆  

Fungus-inhibitory Activity and Gene Cloning of β-glucanase from Bacillus velezensis YB15

作　　者：徐婷[1] 朱天辉[1] 李姝江[2] 谯天敏 

机构地区：[1]四川农业大学林学院,雅安625014 [2]四川农业大学森林保护省级重点实验室,雅安625014 [3]长江上游林业生态工程省级重点实验室,雅安625014

出　　处：《中国生物防治学报》2014年第2期276-281,共6页Chinese Journal of Biological Control

基　　金：国家自然科技资源共享平台(2005DKA21207-13);大学生创新试验计划项目(1310626026);四川省教育厅重点项目(14ZA113)

摘　　要：本文在对峙法验证贝莱斯芽孢杆菌B.velezensis YB15具抑菌作用的基础上,用透明圈法、DNS法研究其产β-葡聚糖酶特性,利用对峙法验证该酶抑菌作用,通过PCR法获得目的基因,分析克隆序列并预测其蛋白质结构与功能。结果表明,该菌株对多种病原真菌有拮抗作用,杨树紫纹羽病菌拮抗带达11.0 mm,该菌提取的葡聚糖酶粗酶液对杨树紫纹羽病菌抑菌带为10.6 mm,说明葡聚糖酶在菌株YB15抑菌中有重要作用。不同接种方法影响菌株YB15葡聚糖酶水解透明圈形成,点种法水解圈与菌落直径之比在72 h可达14.1,效果最好。克隆所得菌株YB15葡聚糖酶基因命名为Bglu1,该基因序列长732 bp,编码243个氨基酸,此酶蛋白氨基酸序列与解淀粉芽孢杆菌TB2β-1,3-1,4-葡聚糖酶同源性较高,属糖基水解酶16家族,N端疏水区存在信号肽并具跨膜区域,推测其为分泌蛋白。On the basis of fungus-inhibitory activity tested with dual culture method, authors studied characteristics of β-glucanase produced by B. velezensis YB15. Results showed that the strain played an antagonistic role against many pathogenic fungi. Width of inhibition zone against Helicobasidium purpureum was 11.0 and 10.6 mm by YB15 and itsβ-glucanase crude enzyme, respectively. Different inoculation methods could impose impacts on the formation of the hydrolytic transparent circle of the glucanase of YB15. The proportion of hydrolytic circle and colony diameter by dibbling could be up to 14.1 at 72 h. The cloned gene of YB15β-glucanase named as Bglu1 was 732 bp in length and encoded 243 amino acids. The protein showed high homology with β-1,3-1,4-glucanase of B. amyloliquefaciens TB2 belonging to glycosyl hydrolase-16 family. The N-terminal hydrophobic domain had signal peptide and transmembrance domain.

关 键 词：贝莱斯芽孢杆菌 Β-葡聚糖酶 抑菌作用 基因克隆 

分 类 号：S828.5[农业科学—畜牧学] TQ925.1[农业科学—畜牧兽医]