DNA磷硫酰化修饰是首次在DNA大分子骨架上发现的表观遗传修饰。该修饰广泛存在于细菌中并具有序列和手性特异性。为了检测大肠杆菌Escherichia coli B7A基因组水平上DNA磷硫酰化修饰位点，发展了单分子实时测序（single molecule real-time sequencing, SMRT）和碘切割依赖的深度测序（deep sequencing of iodine-induced cleavage,ICDS)两种技术。B7A基因组磷硫酰化修饰主要发生在部分的双链的GpsAAC/GpsTTC序列上。本项目在此基础上，拟测定多个细菌基因组磷硫酰化修饰位点，深入开展磷硫酰化修饰的序列多样性和在基因组中的分布研究，解析B7A基因组中GAAC/GTTC位点部分修饰的分子机制，阐明DNA磷硫酰化修饰的序列特征。为揭示DNA磷硫酰化修饰的生物化学机制和生物学功能奠定基础。

DNA phosphorothioations are an epigenetic modification firstly discovered on the DNA backbone, which are widespread in many species of bacteria with stereo-specificity and diverse sequence contexts. To detect the distribution of phosphorothioate modifications across the Escherichia coli B7A genome, two new complementary technologies: single molecule, real-time (SMRT) sequencing and deep sequencing of iodine-induced cleavage at phosphorothioation (ICDS) have been developed, and showed that phosphorothioation was detected mainly in GpsAAC/GpsTTC sequences on both strands in B7A. The sequence diversity, abundance and the distribution of DNA phosphorothioate modification in several different bacterial genomes will be investigated by using both SMRT and ICDS sequencing. The molecular mechanism of partial modification of consensus sequence GAAC/GTTC in E.coli B7A, and the sequence specificity of phosphorothioation consensus sites will be elucidated. The goal of this project is to develop and apply novel sequencing technologies to localize phosphorothioate modifications in bacterial genomes as part of a multifaceted effort to define the biochemical mechanism and the biological function of phosphorothioate modifications.