Timm J, Lim E M, Gicquel B
Unité de Génétique Mycobactérienne, CNRS URA 1300, Institut Pasteur, Paris, France.
J Bacteriol. 1994 Nov;176(21):6749-53. doi: 10.1128/jb.176.21.6749-6753.1994.
A series of Escherichia coli-mycobacteria shuttle plasmids for the isolation and study of gene regulatory sequences was constructed. These pJEM vectors contain an efficient transcription terminator and multiple cloning sites and allow either operon or gene fusions to lacZ. By constructing operon fusions with pJEM15, we assessed various previously characterized mycobacterial promoters in the fast-growing species Mycobacterium smegmatis and the slow-growing species M. bovis BCG. Our results suggest that M. smegmatis and M. bovis BCG RNA polymerases do not share the same specificity. To isolate new mycobacterial promoters, an M. tuberculosis DNA library was generated, using pJEM13, and screened in M. smegmatis. Several Lac+ clones were isolated, and the beta-galactosidase activity was measured.
构建了一系列用于分离和研究基因调控序列的大肠杆菌-分枝杆菌穿梭质粒。这些pJEM载体包含一个高效转录终止子和多个克隆位点，并允许与lacZ进行操纵子或基因融合。通过用pJEM15构建操纵子融合体，我们评估了快速生长的耻垢分枝杆菌和缓慢生长的牛分枝杆菌卡介苗中各种先前已鉴定的分枝杆菌启动子。我们的结果表明，耻垢分枝杆菌和牛分枝杆菌卡介苗的RNA聚合酶不具有相同的特异性。为了分离新的分枝杆菌启动子，使用pJEM13构建了结核分枝杆菌DNA文库，并在耻垢分枝杆菌中进行筛选。分离出了几个Lac+克隆，并测量了β-半乳糖苷酶活性。
