MUTANTS IN BDELLOVIBRIO DEVELOPMENT

蛭弧菌发育中的突变体

• 批准号: 2191751
• 负责人: JOHN J TUDOR
• 金额: $ 11.24万
• 依托单位: SAINT JOSEPH'S UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-01 至 1999-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2191751
• 关键词: Escherichia coli; bacterial DNA; bacterial genetics; bacteriolysis; cell cycle; genetic library; gram negative bacteria; life cycle; microorganism growth; nucleic acid sequence; plaque assay; plasmids; temperature sensitive mutant

DESCRIPTION: Members of the bacterial genus Bdellovibrio are obligate predators of other Gram negative bacteria. They follow a biphasic life cycle with a free swimming attack phase and an intracellular growth phase. Nothing is currently known regarding gene regulation in bdellovibrios, indeed little is known about their genetics in general. However, a system for introducing DNA into bdellovibrios has recently been reported, making some genetic approaches possible in this organism. The investigator proposes to isolate and characterize conditional mutants that have lost the ability to produce plaques on lawns of susceptible prey cells. The overall objectives are to isolate temperature-sensitive (ts) mutants of the developmental cycle of the bdellovibrios, to characterize those mutants, and to isolate and characterize mutant DNA sequences. Bdellovibrios will be chemically mutagenized, and mutants unable to produce plaques at 34 degrees C but able to plaque at 20 degrees C will be isolated. Each mutant isolated will be characterized according to which phase in the developmental cycle is blocked. Mutants unable to penetrate into the prey's periplasmic space will be further examined for altered enzymatic activity of penetration-specific enzymes. Mutants from each category will be mated with E. coli carrying wild-type Bdellovibrio genomic libraries in transmissible vectors. Conjugants that have regained the ability to produce plaques at 34 degrees C will be isolated, plasmid DNA extracted and mutant sequences characterized by restriction analysis and DNA sequencing analysis in comparison to wild-type sequences. Sequences obtained will be screened against known sequences in Genbank for homology. Identification with homologous sequences may suggest possible functions of the gene products involved. This work provides an initial step towards understanding the nature of the Bdellovibrio-prey cell interaction.

产品说明： 蛭弧菌属细菌的成员是专性的， 其他革兰氏阴性菌的捕食者。 他们遵循一种双相生活 具有自由游泳攻击阶段和细胞内生长的周期 相位 目前还不知道基因调控， 事实上，对于蛭弧菌，人们对它们的遗传学知之甚少。 然而，最近将DNA引入蛭弧菌的系统 已经报道，使一些遗传方法在这种生物体中成为可能。 研究者建议分离和表征条件性 突变体已经失去了在草坪上产生斑块的能力， 易感的猎物细胞 总体目标是分离 温度敏感（TS）突变体的发育周期的 蛭弧菌，以表征这些突变体，并分离和 表征突变DNA序列。 蛭弧菌会在化学上 突变，突变体不能在34摄氏度产生斑块， 能够在20摄氏度的斑块将被隔离。 每个分离的突变体 将根据发展的哪个阶段来描述 循环被阻塞。 变种人无法穿透猎物的 将进一步检查周质间隙中酶的改变 渗透特异性酶的活性。 每个类别的突变体 将与E交配。携带野生型蛭弧菌基因组 可传播载体中的文库。 已经恢复了免疫原性的缀合物 能够在34摄氏度产生噬斑将被分离，质粒DNA 通过限制性分析表征的提取的和突变的序列， 与野生型序列相比的DNA测序分析。 序列 将针对Genbank中的已知序列筛选获得的基因， 同源性 同源序列的鉴定可能表明 基因产物的功能。 这项工作提供了一个初步的 进一步了解蛭弧菌-猎物细胞的性质 互动