Kormanec J, Sevcíková B, Sprusanský O, Benada O, Kofronová O, Nováková R, Rezuchová B, Potúcková L, Homérová D
Institute of Molecular Biology, Slovak Academy of Sciences, Bratislava, Slovak Republic.
Folia Microbiol (Praha). 1998;43(6):605-12. doi: 10.1007/BF02816376.
In previous experiments, a Streptomyces aureofaciens gene highly similar to the sporulation-specific whiB gene of Streptomyces coelicolor was identified. By integrative transformation, via double cross-over, a stable null mutant of the whiB-homologous gene of S. aureofaciens was obtained. The disruption blocked differentiation at a stage between the formation of aerial mycelium and the development of mature spores, producing white aerial hyphae without septation. Expression of the whiB gene was investigated during differentiation by S1 nuclease mapping, using RNA prepared from S. aureofaciens in various developmental stages. Two putative promoters were identified upstream of the whiB coding region. The stronger promoter, whiB-P2, was induced at the beginning of aerial mycelium formation, and the weaker promoter, whiB-P1, was expressed fairly constantly during differentiation. No differences in the expression of the whiB promoters were detected in an rpoZ-disrupted S. aureofaciens strain. The promoter bearing DNA fragment was inserted into the promoter-probe vector pARC1 to produce an expression pattern consistent with the results of direct RNA analysis.
在先前的实验中,鉴定出了一个与天蓝色链霉菌的孢子形成特异性whiB基因高度相似的金色链霉菌基因。通过整合转化,经双交换获得了金色链霉菌whiB同源基因的稳定无效突变体。该破坏作用在气生菌丝形成和成熟孢子发育之间的阶段阻断了分化,产生了无隔膜的白色气生菌丝。利用从处于不同发育阶段的金色链霉菌制备的RNA,通过S1核酸酶作图研究了whiB基因在分化过程中的表达。在whiB编码区上游鉴定出两个推定的启动子。较强的启动子whiB-P2在气生菌丝形成开始时被诱导,较弱的启动子whiB-P1在分化过程中表达相当稳定。在rpoZ基因被破坏的金色链霉菌菌株中未检测到whiB启动子表达的差异。将携带启动子的DNA片段插入启动子探针载体pARC1中,以产生与直接RNA分析结果一致的表达模式。