Shimizu T, Kobayashi T, Ba-Thein W, Ohtani K, Hayashi H
Department of Microbiology, University of Tsukuba, Ibaraki, Japan.
Microbiol Immunol. 1995;39(9):677-86. doi: 10.1111/j.1348-0421.1995.tb03256.x.
The 3'-flanking region of the perfringolysin O (theta-toxin) gene (pfoA) of Clostridium perfringens was analyzed by chromosome walking. A total of 5,363 bp of the downstream region of the pfoA gene was sequenced and four open reading frames were found. ORF54 and ORF80 were found to be homologous to genes coding for membrane-bound transporter proteins of other bacteria and the beta-galactosidase gene (bgaB) of Bacillus stearothermophilus, respectively. ORF80 was named the pbg gene. Clones which showed beta-galactosidase activities were selected from a lambda FIXII genomic library of C. perfringens by blue plaque screening using X-Gal as a substrate. Four clones whose plaques showed blue appearances were obtained. Two of the four clones hybridized with the pbg probe but the others did not, indicating that there are two distinct beta-galactosidase genes in C. perfringens. The pbg gene was subcloned into pBR322 and was successfully expressed in Escherichia coli, suggesting that the pbg gene codes for a beta-galactosidase of C. perfringens.
通过染色体步移技术对产气荚膜梭菌的穿孔毒素O（θ毒素）基因（pfoA）的3'侧翼区域进行了分析。对pfoA基因下游区域总共5363 bp进行了测序，发现了四个开放阅读框。发现ORF54和ORF80分别与编码其他细菌膜结合转运蛋白的基因以及嗜热脂肪芽孢杆菌的β-半乳糖苷酶基因（bgaB）同源。将ORF80命名为pbg基因。以X-Gal为底物通过蓝斑筛选从产气荚膜梭菌的λFIXII基因组文库中筛选出具有β-半乳糖苷酶活性的克隆。获得了四个菌斑呈蓝色的克隆。四个克隆中有两个与pbg探针杂交，而其他克隆则没有，这表明产气荚膜梭菌中有两个不同的β-半乳糖苷酶基因。将pbg基因亚克隆到pBR322中，并在大肠杆菌中成功表达，这表明pbg基因编码产气荚膜梭菌的一种β-半乳糖苷酶。
