创建针对调控元件的基因编辑工具是研究非编码序列功能的重要前提。基于CRISPR的碱基编辑技术，因其碱基原位替换的编辑优势，成为研究非编码区调控元件的有利工具。由于调控元件由多碱基构成，针对单碱基突变通常不足以改变其功能，申请人前期构建了BE-PLUS，具备宽窗口特性，可经多碱基饱和突变破坏调控元件，更适于针对调控元件的研究。然而调控区域富含“GC”序列，BE-PLUS中的脱氨酶rAPOBEC1在“GC”序列中效率低，应用受限。已知的脱氨酶AID虽无明显序列偏好性，但编辑效率不足以满足需求。本研究拟采用密码子优化、截短的脱氨酶hAID替代BE-PLUS中的rAPOBEC1，建立新型碱基编辑工具BE-PLUS-AID，并通过编辑DNA调控元件G-四联体进行验证，完成序列兼容性、宽窗口的高效碱基编辑工具的构建，并应用于非编码区调控元件。本课题对基因组中非编码区调控元件的功能研究具有重要科学意义。

Creating a genome editing tool for regulatory elements is an important prerequisite for the functional research of non-coding sequences. CRISPR-mediated base editing technology induces base substitutions in situ, making it a useful tool for studying the non-coding regulatory elements. Regulatory element is normally composed of multiple bases, in which single base mutation is usually inadequate to affect its function. Constructed previously by the applicant, BE-PLUS is a suitable tool to study regulatory elements for its ability to destroy the regulatory element by multi-base mutagenesis with its wide window. However, because of its deaminase rAPOBEC1 is inefficient in “GC” sequences, BE-PLUS is limited in its application to study “GC” sequence enriched regulatory regions. Deaminase AID has no significant sequence preferences, but not efficient enough. Here we optimized BE-PLUS system by replacing the cytidine deaminase rAPOBEC1 with codon optimized truncate hAID to construct an advanced base editor, called BE-PLUS-AID. Next, we will verify its function by editing DNA G-quadruplex sequences in regulation regions, achieving broad window and efficient base editing with sequence compatibility in non-coding regulatory elements. If verified, BE-PLUS-AID could make a great contribution in the investigation of non-coding regulatory elements.