| 具分泌几丁质酶活性的生防细菌的筛选鉴定及其几丁质酶基因的克隆和表达 |
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| 引用本文： | 石磊,,杜锦锦,郭庆港,李宝庆,鹿秀云,李社增,马平.具分泌几丁质酶活性的生防细菌的筛选鉴定及其几丁质酶基因的克隆和表达[J].植物病理学报,2013,43(2):149-156. |
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| 作者姓名： | 石磊 杜锦锦 郭庆港 李宝庆 鹿秀云 李社增 马平 |
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| 作者单位： | 1、 河北省农林科学院植物保护研究所, 河北省农业有害生物综合防治工程技术研究中心,农业部华北北部作物有害生物综合治理重点实验室,保定 071000; 2、 河北农业大学植物保护学院,保定 071000 |
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| 基金项目： | 国家高技术研究发展计划（863计划）（2011AA10A205）;河北省财政专项（F12C10036） |
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| 摘 要： | 本研究通过平板透明圈法筛选获得一株具有几丁质酶活性的生防细菌CAB-1,该菌株对番茄灰霉病菌等多种病原真菌表现较强的拮抗活性。通过生理生化、16S rDNA和gyrB基因序列测定,将菌株CAB-1鉴定为萎缩芽胞杆菌（Bacillus atrophaeus）。对菌株CAB-1全基因组序列进行分析和功能预测,发现该菌株存在2个几丁质酶编码基因chit1和chit2。通过PCR技术从菌株CAB-1中克隆出这两个几丁质酶的编码基因并在大肠杆菌中表达,其原核表达产物均表现几丁质酶活性,其中chit1的原核表达产物能够显著抑制灰霉菌分生孢子的萌发。对其原核表达条件进行优化,发现在30℃下振荡培养24 h,IPTG浓度为0.2~1.0 mmol/L时,其蛋白表达量最高。
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| 关 键 词： | 萎缩芽胞杆菌 生防菌 几丁质酶 基因克隆 原核表达 |
| 收稿时间： | 2012-07-25; |
Screening and identification of an antifungal chitinolytic bacterium and its gene cloning and expressing |
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| SHI Lei,,DU Jin-jin,GUO Qing-gang,LI Bao-qing,LU Xiu-yun,LI She-zeng,MA Ping.Screening and identification of an antifungal chitinolytic bacterium and its gene cloning and expressing[J].Acta Phytopathologica Sinica,2013,43(2):149-156. |
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| Authors: | SHI Lei DU Jin-jin GUO Qing-gang LI Bao-qing LU Xiu-yun LI She-zeng MA Ping |
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| Institution: | 1.Plant Protection Institute Hebei Academy of Agriculture and Forestry Sciences, IPM centre of Hebei Province, Key Laboratory of IPM on Crops in Northern Region of North China, Ministry of Agriculture, Baoding 071000, China; 2.College of Plant Protection, Agricultural University of Hebei, Baoding 071000, China |
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| Abstract: | Bacterial strain CAB-1 was isolated according to its chitinolytic activity and antifungal activities against plant pathogens including Botrytis cinerea in virto. Strain CAB-1 was identified as Bacillus atrophaeus based on the biochemical characters, sequences analysis of 16S rDNA and gyrB. Through sequence analysis and function predicting based on it whole genome, two chitinase genes were located and cloned from strain CAB-1. These two cloned chitinase genes were ligated into the prokaryotic expression vector and fusion proteins were induced. Chitinase activities of the induced protein Chit1 and Chit2 were testified by plate transparent circle method. In the meantime, only Chit1 protein showed antifungal activity against B. cinerea. The expression condition of Chit1 protein was optimized. The results showed that incubating at 30℃ for 24 hours in shaker and induced by IPTG of 0.2 mmol/L to 1.0 mmol/L was recognized as the optimized condition for Chit1 protein production. |
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| Keywords: | Bacillus atrophaeus Bacillus atrophaeus biocontrol bacteria chitinase gene cloning prokaryotic expression |
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