Picardeau Mathieu, Le Dantec Corinne, Vincent Véronique
Laboratoire de Référence des Mycobactéries, Institut Pasteur, 75724 Paris Cedex 15, France1.
Microbiology (Reading). 2000 Feb;146 ( Pt 2):305-313. doi: 10.1099/00221287-146-2-305.
Linear plasmids have previously been identified by the authors in mycobacteria, the telomeres of which have terminal inverted repeats and covalently attached proteins. In this study, the replication of these unusual molecules was investigated by studying a 25 kb linear plasmid from the slow-growing species Mycobacterium celatum called pCLP. An internal region of pCLP responsible for replication in Mycobacterium smegmatis was identified. The nucleotide sequence of the minimum replication region of pCLP, which was 2.8 kb long, contained a putative replication gene, rep, and a putative origin of replication consisting of an 18 bp direct repeat and an AT-rich region. A short section of the pCLP replication region was also found to have sequence identity with the replication regions of mycobacterial circular plasmids, suggesting that these linear and circular plasmids are related. It was found that pCLP replicated in Mycobacterium bovis BCG and was compatible in M. smegmatis with pAL5000- and pJAZ38-derived plasmids from Mycobacterium fortuitum, which belong to two different compatibility groups. Thus, this new Escherichia coli-mycobacteria shuttle vector may be used in both slow- and fast-growing mycobacteria and in co-transformation experiments with other mycobacterial vectors.
作者先前已在分枝杆菌中鉴定出线性质粒，其端粒具有末端反向重复序列和共价连接的蛋白质。在本研究中，通过研究来自生长缓慢的塞拉分枝杆菌的一个25 kb线性质粒pCLP，对这些不同寻常的分子的复制进行了研究。确定了pCLP在耻垢分枝杆菌中负责复制的内部区域。pCLP最小复制区域的核苷酸序列长2.8 kb，包含一个假定的复制基因rep和一个由18 bp直接重复序列和富含AT区域组成的假定复制起点。还发现pCLP复制区域的一小段与分枝杆菌环状质粒的复制区域具有序列同一性，这表明这些线性和环状质粒是相关的。研究发现pCLP能在牛分枝杆菌卡介苗中复制，并且在耻垢分枝杆菌中与来自偶然分枝杆菌的pAL5000和pJAZ38衍生质粒兼容，这两种质粒属于两个不同的相容性群。因此，这种新型大肠杆菌-分枝杆菌穿梭载体可用于生长缓慢和快速生长的分枝杆菌，以及与其他分枝杆菌载体的共转化实验。
