单细胞RNA m5C测序技术研发

本项目切合指南重点资助研究方向（一）“生物大分子动态修饰的化学标记与检测技术”。单细胞测序技术已经成为分析细胞异质性、细胞分化及发育的重要技术手段，而表观遗传修饰在细胞异质性调控方面发挥重要作用。单细胞DNA m5C甲基化测序技术已被研发，但尚未实现单细胞RNA m5C甲基化测序技术。前期研究中，申请人建立了单细胞总RNA测序新技术，优化了单细胞RNA m5C修饰测序技术流程，探索了单细胞分选、亚硫酸氢盐处理及文库构建方法，优化了组织和胚胎单细胞水平RNA m5C修饰鉴定流程。本项目利用化学生物学、分子生物学、生物化学、基因组学与生物信息学等交叉学科技术，旨在建立单细胞总RNA测序新技术及单细胞RNA m5C修饰测序技术，绘制单细胞RNA m5C修饰图谱，探究单细胞RNA m5C修饰在斑马鱼胚胎发育中的生物学功能，为阐明单细胞RNA m5C修饰表观遗传异质性研究提供技术支持。

This project is in line with the key research direction of the guide (I) "chemical labeling and detection technology of dynamic modification of biomacromolecules". Single cell sequencing has become an important technique to analyze cell heterogeneity, cell differentiation and development. Epigenetic modifications play important roles in the regulation of cell heterogeneity. Single cell DNA m5C sequencing technology has been developed, but single cell RNA m5C sequencing technology has not been realized. In the preliminary study, we have established a new technology of single cell total RNA sequencing, optimized the single cell RNA m5C sequencing technology principle, explored the methods for single cell sorting, bisulfite treatment and library construction, and optimized RNA m5C identification process in the single cell level of tissue and embryo. By integrating multi-cross-disciplinary technologies, including chemical biology, molecular biology, biochemistry, genomics and bioinformatics, this project aims to establish new technology of single cell total RNA sequencing and single cell RNA m5C sequencing, draw single cell RNA m5C modification landscape, identify the biological functions of single cell RNA m5C in zebrafish embryonic development. This study will provide technical support for clarifying the epigenetic heterogeneity of single cell RNA m5C modification.