Genome wide binding sites of NsrR in uropathogenic E.coli CFT073

We applied ChIP-seq to identify genome wide binding targets of NsrR in E.coli CFT073. NsrR is a nitric oxide sensitive regulator of transcription. Genome wide binding targets of NsrR have been identified in E.coli K12 using ChIP-chip. The genome of CFT073 is about 0.6Mb larger than that of K12. In this study, we identify the novel NsrR binding sites in CFT073. The nsrR gene was modified by the addition of DNA sequences encoding a C-terminal 3X-Flag tag, and the tagged gene was integrated into the chromosome. NsrR bound DNA was isolated by chromatin immunoprecipitation and it was sequenced using Miseq platform.

本研究采用染色质免疫共沉淀测序(Chromatin Immunoprecipitation Sequencing, ChIP-seq)技术，鉴定大肠杆菌CFT073(Escherichia coli CFT073)中NsrR的全基因组结合靶标。NsrR是一类对一氧化氮敏感的转录调节因子。此前已有研究通过染色质免疫共沉淀芯片(Chromatin Immunoprecipitation on Chip, ChIP-chip)在大肠杆菌K12(Escherichia coli K12)中鉴定出NsrR的全基因组结合靶标。大肠杆菌CFT073的基因组较K12大约0.6兆碱基。本研究旨在鉴定CFT073中全新的NsrR结合位点：我们对nsrR基因进行修饰，在其编码区C端添加编码三拷贝Flag(3X-Flag)标签的DNA序列，并将该带标签的基因整合至细菌染色体中。通过染色质免疫共沉淀分离得到结合NsrR的DNA片段，随后采用Miseq测序平台完成测序。