USE OF CHROMOSOMAL INTEGRATION IN THE ESTABLISHMENT AND EXPRESSION OF BLAZ, A STAPHYLOCOCCUS-AUREUS BETA-LACTAMASE GENE, IN BACILLUS-SUBTILIS
USE OF CHROMOSOMAL INTEGRATION IN THE ESTABLISHMENT AND EXPRESSION OF BLAZ, A STAPHYLOCOCCUS-AUREUS BETA-LACTAMASE GENE, IN BACILLUS-SUBTILIS
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DOI：
10.1128/jb.157.3.718-726.1984
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发表时间：
1984-01-01
影响因子：
3.2
通讯作者：
GUYER, MS
中科院分区：
文献类型：
--
作者：
SAUNDERS, CW;SCHMIDT, BJ;GUYER, MS
利用几种不同的载体，尝试在枯草杆菌中建立一种金黄色葡萄球菌β-内酰胺酶基因Blaz。BLAZ基因在枯草杆菌中的稳定建立是通过使用一种载体实现的，该载体允许将该基因的一个拷贝整合到该宿主的染色体中。BLAZ在异源宿主中的表达是因为携带整合BLAZ的枯草杆菌产生β-内酰胺酶，并且比野生型枯草芽孢杆菌对氨苄西林的抗性更强。BLAZ在这些菌株中稳定遗传，因为在非选择性液体培养基或固体培养基上生长后没有观察到产生β-内酰胺酶的能力丧失。以pUB110或pC194为基础的载体不能在枯草杆菌中建立含有BLAZ的限制性片段。同样，含有BLAZ 5‘端(包括信号序列的启动子和编码区以及成熟蛋白的前几个氨基酸)的pC194穿梭载体(PGX318)不能转化枯草杆菌。分离到两个能在枯草杆菌中稳定存在的pGX318衍生物。这些衍生物的结构表明，Blaz启动子的失活与获得待建立的能力有关。
With several different vectors, attempts were made to establish blaZ, a S. aureus .beta.-lactamase gene, in B. subtilis. Stable establishment of blaZ in B. subtilis was achieved by use of a vector that allowed the integration of a single copy of the gene into the chromosome of that host. blaZ was expressed in the heterologous host since B. subtilis strains carrying integrated blaZ produced .beta.-lactamase and were more resistant to ampicillin than was wild-type B. subtilis. blaZ was stably inherited in such strains, as no loss of the ability to produce .beta.-lactamase was observed after growth in nonselective liquid medium or on solid medium. A blaZ-containing restriction fragment could not be established in B. subtilis with either pUB110- or pC194-based vectors. Similarly, a pC194-based shuttle vector (pGX318) containing the 5'' end of blaZ (including the promoter and the coding region for the signal sequence and the first few amino acids of the mature protein) was unable to transform B. subtilis. Two derivatives of pGX318 that could be stably established in B. subtilis were isolated. The structures of these derivatives suggested that inactivation of the blaZ promoter was associated with the acquisition of the ability to be established.