Control of Heterocyst Differentiation in Free-Living and Symbiotic Cyanobacteria

PI假设调控异形胞分化的机制(发菜的好氧固氮部位)。当发菜处于共生状态时，与自由生长状态下的生长不同。他计划使用生理学和分子遗传学，在自由生活和共生生长状态下进行分析，以检验这一假说。现有的突变体将在体外连接到质粒后通过基因组DNA的电穿孔来补充。插入突变将被用来分离更多的突变体。中断的基因将通过回收插入元件来分离；与插入片段相邻的基因组DNA将从粘粒文库中克隆并进行鉴定。葡萄糖-6-磷酸脱氢酶(ZWF)的转录调控将利用最近克隆的基因来分析氮剥夺和共生相互作用的响应。我们将利用zwf的插入突变体来分析其在杂胞体功能中的作用。最近发现的潜在的氮和厌氧调节基因将通过异源杂交从粘粒文库中分离出来，并用于构建发菜的插入突变体，用于分析异形胞的分化。这些结果将有助于理解丝状蓝藻在两种不同生长状态下的环境信号和细胞分化调节机制。

The PI hypothesizes that the mechanisms regulating the differentiation of heterocysts (sites of aerobic N2 fixation by Nostoc sp. differ when Nostoc is in symbiotic association relative to growth in the free-living state. He plans to use physiological and molecular genetics to test the hypothesis with analyses in both free-living and symbiotic growth states. Existing mutants will be complemented by electroporation of genomic DNA after in vitro ligation into a plasmid. Insertional mutagenesis will be used to isolate additional mutants. Interrupted genes will be isolated by rescue of the insertion elements; genomic DNA adjacent to the insertions will be cloned form cosmid libraries and characterized. Transcriptional regulation of glucose-6-phosphate dehydrogenase (zwf) will be analyzed in response to nitrogen deprivation and symbiotic interaction using the recently cloned gene. Insertional mutants of zwf will be used to analyze its role in heterocyst function. Recently identified potential nitrogen and anaerobic regulatory genes will be isolated from cosmid libraries by heterologous hybridization and used to construct insertional mutants in Nostoc for analysis of heterocyst differentiation. The results will lead to an understanding of the environmental signals and mechanisms of regulation of cell differentiation in filamentous cyanobacteria in two different growth states.