产微球茎菌琼胶酶基因的克隆及生物信息学分析

以琼脂为唯一碳源的培养基分离出一株产琼胶酶的海洋菌株AG1，16S rRNA基因序列分析显示，该菌株为产微球茎菌（Microbulbifer sp.）。以菌株AG1的基因组为模板，使用琼胶酶特异性引物进行PCR扩增，将扩增产物克隆至pMD18-T载体后进行测序。结果显示，克隆基因的大小为1302 bp，预测编码含有433个氨基酸残基的蛋白质。对该蛋白质进行生物信息学分析，结果表明，该蛋白质序列与来自耐热微泡菌（Microbulbifer thermotolerans）的琼胶酶氨基酸序列相似性为100%，预测本研究克隆的基因编码琼胶酶。该琼胶酶的理论分子质量大小为48.2 ku，理论等电点为5.42。采用同源建模法建立Microbulbifer sp.AG1琼胶酶的三维结构，富含β-折叠。

Cloning and Bioinformatics Analysis of the Agarase Gene from Microbulbifer sp.AG1

By using agar as the sole carbon source in the culture medium,a marine bacterium strain AG1 producing agarases was isolated.It was identified as Microbulbifer sp.AG1 based on 16S rRNA gene sequences alignment.The genomic DNA of this strain was used as the template for amplification of agarase gene by using PCR with a pair of specific primers.The amplified products were cloned into pMD18-T vector and then sequenced.It showed that the cloned gene was 1302 bp, encoding 433 amino acid residues.This protein was further analyzed by bioinformatics.The results showed that the target protein sequence shared 100% identity with the agarase sequence from Microbulbifer thermotolerans.The theoretical molecular weight and pI of the agarase were 48.2 ku and 5.42, respectively.The three-dimensional structure of Microbulbifer sp.AG1 agarase was constructed by homology modeling and presented β-strands rich structure.