林 毅1,2, 彭 锟1,2, 关 雄1**
（1.生物农药与化学生物学教育部重点实验室，福建 福州，350002；2.华侨大学生物工程与技术系，福建 泉州，362021）
摘 要: 从苏云金杆菌（Bacillus thuringiensis, Bt）entomocidus亚种HD109菌株中克隆了几丁质酶基因chiA74-HD109，其序列全长为2031 bp，编码676个氨基酸，GenBank登录号为AY455290。其编码产物与蜡状芽孢杆菌几丁质酶CHiB（AB041932）的相似性达97.9%，与Bt kenyae亚种LBIT-82菌株几丁质酶chiA74 (AF424979) 的相似性达98.4%，与Bt pakistani亚种几丁质酶(U89796)的相似性为83.0%，与Bt kurstaki亚种几丁质酶kchi (AY189740)的相似性达97.8%，与Bt israelensis亚种几丁质酶(AF526379)的相似性达96.6%，与Bt几丁质酶(AY074882)的相似性达98.4%，与Bt sotto亚种几丁质酶(AY129671)的相似性为97.3%。功能结构域分析显示其编码区包含信号肽、催化区、Ⅲ型粘蛋白同源区及几丁质结合区4个部分。
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Cloning and bioinformatics analysis of the chitinase gene from Bacillus thuringiensis serovar entomocidus strain HD109
LIN Yi1,2, PENG Kun1,2, GUAN Xiong1**
(1. Key Lab of Biopesticide and chemical biology, Ministry of Education, Fujian Agriculture and Forestry University, Fujian Fuzhou, 350002; 2.Department of Bioengineering and Biotechnology, Huaqiao University, Fujian Quanzhou, 362021)
Abstract:Based on the conserved 5' and 3' sequence of chitinase gene from B.thuringiensis, a product of 2031 bp was amplified and cloned into Escherichia coli strain DH5α from B.thuringiensis serovar entomocidus strain HD109 genomic DNA. The product encoded an open reading frame (chiA74-HD109) encoding a deduced protein of 676 amino acids. Removal of the signal peptide sequence resulted in a predicted protein that was 70457 Da in size and 5.45 of isoelectric point. The deduced 676 amino acids sequence showed high degree of identity with other chitinases such as ChiB (AB041932) from Bacillus cereus (97.9%), chiA74 (AF424979) from B.thuringiensis serovar kenyae strain LBIT-82 (98.4%), chitinase (U89796) from B.thuringiensis serovar pakistani (83.0%), kchi (AY189740) from B.thuringiensis serovar kurstaki (97.8%), chitinase (AF526379) from B.thuringiensis serovar israelensis (96.6%), chitinase (AY074882) from B.thuringiensis (98.4%), and chitinase (AY129671) from B.thuringiensis serovar sotto (97.3%).Analysis of the sequence indicated that the chitinase contained a catalytic domain belonging to family 18 of glycosyl hydrolases in the N-terminus, a fibronectin type Ⅲ domain in the middle region and a chitin-binding domain in the C-terminus.All three domains showed conserved sequences when compared to other bacterial chitinase sequences.
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