Mapping Chromosome Accessability to Transposition Using Microarrays

使用微阵列绘制染色体易接近性至转座的图谱

• 批准号: 0615953
• 负责人: Norman Higgins
• 金额: --
• 依托单位: University of Alabama at Birmingham
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0615953&HistoricalAwards=false
• 关键词: Mapping; Chromosome; Accessability; Transposition; Using

Two related methods have been developed to study chromosome structure inside living bacteria. One technique called Muprinting uses PCR reactions to generate high resolution maps of phage Mu insertions in chromosomes. By comparing in vitro and in vivo Muprints of specific DNA regions, the binding of repressors, chromosome partitioning proteins, and RNA polymerase can be monitored. A new method exploits DNA microarrays and bioinformatics to generate genome-wide scans of chromosome structure. Microarray "snapshots" identify chromosomal locations with very high and very low frequencies of transposon insertions. Three goals of the current project are: (i) Make genome snapshots of chromosomes for cells growing with 90 min. doubling time (minimal glucose medium), 125 min doubling time (minimal alanine) , and 300 min. doubling time (minimal succinate medium). When combined with snapshots from cells growing at maximal speed, this information shows how chromosome structure changes in response to cellular metabolic needs. (ii) Define molecular mechanisms that enhance and cloak DNA accessibility to Mu transposition. DNA sites where cis- and trans-regulatory factors are implicated in genetic control will be studied using high resolution in vitro and in vivo Muprint reactions. (iii) E. coli and S. typhimurium transposition patterns will be compared to learn how evolution has altered transposon in the 63% of genes that have been conserved since divergence of these two bacteria from common ancestors. These studies provide a new biochemical model to understand chromosome structure and allow students from high school to graduate school to study evolutionary differences in two genetically related bacteria.

已经开发了两种相关的方法来研究活细菌内部的染色体结构。一种称为muprinting的技术使用PCR反应来生成染色体中噬菌体MU插入的高分辨率图。通过比较特定DNA区域的体外和体内默认，可以监测阻遏物的结合，染色体分配蛋白和RNA聚合酶的结合。一种新方法利用了DNA微阵列和生物信息学来生成全基因组结构的扫描。微阵列“快照”识别具有非常高和非常低频率转座子插入的染色体位置。当前项目的三个目标是：（i）制作生长90分钟的细胞染色体的基因组快照。加倍时间（最小葡萄糖培养基），125分钟的两倍时间（最小丙氨酸）和300分钟。两倍时间（最小的琥珀酸酯培养基）。当与以最大速度生长的细胞生长的快照相结合时，该信息显示了染色体结构如何响应细胞代谢需求而变化。 （ii）定义分子机制，从而增强和掩护DNA访问MU转座的可及性。将使用高分辨率的体外和体内默认反应来研究与遗传控制有关的DNA位点。 （iii）将大肠杆菌和鼠伤寒沙门氏菌换位模式进行比较，以了解以来的63％基因中的转化如何改变了这两种细菌与共同祖先的差异以来一直保守的基因。这些研究提供了一种新的生化模型，以了解染色体结构，并允许高中的学生能够研究两种遗传学细菌的进化差异。