细菌DNA磷硫酰化修饰位点的特异性标记

DNA phosphorothioate (PT) modification is a sulfur modification on DNA backbone, in which a non-bridging oxygen is replaced by sulfur in a sequence-selective and Rp-specific manner. As a novel restriction-modification system, DNA PT modification might play a role similar to that of DNA methylation in protecting host's DNA from foreign DNA invasion. Currently, little is known about this novel DNA modification, including its biochemical properties, physiological functions, especially the intuitive labeling of PT site in genome DNA and many other aspects. For the investigation of the modification mechanism and function, site distribution about the PT modification is thus important. In this study, we plan to selectively target the PT modifications in bacterial DNA with small molecules, quantum dots (QDs) or Au anoparticles (AuNPs). Using this method, PT locations in bacterial DNA would be specifically detected by SMRT, Super-Resolution Fluorescence Microscopy and AFM, respectively. Collectively, this study will provide new insights into the distribution and modification frequency characteristics of this new DNA modification.

DNA磷硫酰化修饰是指在天然的DNA磷酸骨架上非桥联氧原子被硫原子所取代的生理修饰，并且这种磷硫酰化修饰具有序列选择性和Rp空间构象专一性的特点。DNA磷硫酰化修饰与DNA甲基化修饰系统类似，属于一种新型的限制-修饰防御系统，具有保护自身DNA免受损伤和维持自身遗传稳定性的作用。DNA磷硫酰化修饰作为一个新的研究领域，人们对这一新型DNA修饰仍然知之甚少，包括修饰的生物化学特性，生理学功能，尤其是在基因组上分布的直观标记等诸多方面。为了更好地探索DNA磷硫酰化修饰机制与功能，本项目拟利用硫原子特征反应，对DNA磷硫酰化修饰位点使用小分子化合物、量子点和纳米金颗粒进行特异性标记，然后分别借助SMRT技术、超分辨率荧光显微镜和原子力显微镜对细菌DNA磷硫酰化修饰位点进行鉴定，揭示这种新型DNA修饰的修饰位点分布和修饰频率特征。

在细菌DNA中发现磷硫酰化 (PT) 修饰，其中硫取代了DNA骨架中的非桥接氧，挑战了我们对细胞DNA保守骨架结构的理解。目前，除了其生化特性外，对这种新型DNA修饰知之甚少，尤其是基因组DNA中的PT位点。因此，为了研究该修饰系统的机制和功能，PT修饰位点的分布和定量信息尤其重要。在我们的研究工作中，细菌DNA中的PT修饰位点被荧光纳米粒子选择性靶向标记，然后进行DNA拉伸和固定。这将为这种新型DNA修饰的分布和修饰频率特征，以及人类基因组精细绘制和疾病相关基因检测和量化的潜在应用提供新的见解。