Cloning of Genes Transcribed by Alternate Sigma Factors

9316114 Jonas该项目将导致革兰氏阳性原核生物枯草芽孢杆菌中交替sigma因子转录的基因的鉴定。 具有被特定σ因子识别的启动子的基因将使用转座子来鉴定，所述转座子经工程改造以包含报告基因（Tn917 lac），其应在B的染色体中产生随机插入突变（和转录融合）。枯草杆菌。 将在B菌株中筛选这些突变。含有B活性形式的结构基因的诱导拷贝（在spac启动子的控制下）的枯草杆菌。或大肠在这样的系统中由IPTG诱导的带有插入的基因将被定位、克隆和测序。 这些基因的启动子区将进行分析，提供有关的信息，由替代西格玛因子使用的调控序列。 这些基因的功能可以通过插入突变体的表型或与已知功能基因的序列同源性来揭示。 我们将试图通过识别调节子中基因的功能来确定替代性σ因子在体内所起的作用。 获得的关于遗传调控的知识可能证明对发酵工业有用，其可以受益于使用替代西格玛因子在细菌中过量产生基因产物。 ***

9316114 Jonas This project will result in the identification of genes transcribed by alternate sigma factors in the Gram positive prokaryote, Bacillus subtilis. Genes having promoters recognized by a particular sigma factor will be identified using a transposon engineered to contain a reporter gene (Tn917lac), which should generate random insertional mutations (and transcriptional fusions) in the chromosome of B. subtilis. These mutations will be screened in strains of B. subtilis containing inducible copies (under control of the spac promoter) of the structural genes for the active forms of B. or E. genes bearing insertion that are induced by IPTG in such a system will be mapped, cloned, and sequenced. The promoter regions of these genes will be analyzed, providing information concerning regulatory sequences used by alternative sigma factors. The functions of these genes may be revealed by the phenotype of the insertional mutants, or by homology with the sequences of genes with known functions. We will attempt to determine the role the alternative sigma factor plays in vivo by recognition of the function of the genes in the regulon. %%% The knowledge gained concerning genetic regulation may prove useful to the fermentation industry, which could benefit from using alternate sigma factors in over producing gene product in bacteria. ***