Regulation of gene expression revealed by genome editing of repetitive and polymorphic cis-elements

通过重复和多态顺式元件的基因组编辑揭示基因表达的调控

• 批准号: 21F21382
• 负责人: ウォルツェン クヌート
• 金额: $ 0.96万
• 依托单位: Kyoto University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for JSPS Fellows
• 财政年份: 2021
• 资助国家: 日本
• 起止时间: 2021-11-18 至 2023-03-31
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-21F21382/
• 关键词: ゲノム編集; ゲノム解析; DNAリピート; ヒトiPS細胞; genome editing; genome analysis; DNA repeat; human iPS cells

The project aims to identify variable tandem repeats (VNTRs) at potential regulatory features, like promoters and enhancers, to inform functional analyses in human iPSCs by gene editing. The candidate developed a computational pipeline to detect tandem repeats at candidate enhancer regions (cCREs) on the ENCODE dataset. The candidate also identified VNTRs in the literature which are proposed to be involved in human disease. The project aims to develop gene editing methods to generate variants in healthy iPSCs. The candidate succeeded in editing repeat contractions at two loci in iPSCs: ADGRG1 (reducing two copies to one copy) and TRIB3 (reducing three or five copies to one copy). TRIB3 editing results have been sequenced using Nanopore long-read technology and an analysis pipeline to genotype the editing results is being established. Moreover, the candidate developed a method based on a novel formulation of gene editing reagents which could be used to generate intermediate numbers of DNA repeat variants. These achievements lay the groundwork for a streamlined pipeline for the analysis of the rest of the identified gene editing sites, enabling the subsequent goals of the project.

该项目旨在识别潜在调控特征（如启动子和增强子）的可变串联重复序列（VNTR），以便通过基因编辑对人类 iPSC 进行功能分析。候选人开发了一个计算管道来检测 ENCODE 数据集上候选增强子区域 (cCRE) 的串联重复。该候选人还在文献中发现了被认为与人类疾病有关的 VNTR。该项目旨在开发基因编辑方法以在健康的 iPSC 中产生变异。该候选者成功编辑了 iPSC 中两个基因座的重复收缩：ADGRG1（将两个副本减少为一个副本）和 TRIB3（将三个或五个副本减少为一个副本）。 TRIB3编辑结果已使用Nanopore长读长技术进行测序，并且正在建立对编辑结果进行基因分型的分析管道。此外，该候选人开发了一种基于基因编辑试剂新配方的方法，可用于生成中间数量的 DNA 重复变异体。这些成就为分析其余已识别基因编辑位点的简化流程奠定了基础，从而实现了该项目的后续目标。

项目成果

Deploying MENdel MMEJ prediction for the efficient generation of frameshift deletions and disease modeling in human induced pluripotent stem cells

部署 MENdel MMEJ 预测以在人类诱导多能干细胞中有效生成移码缺失和疾病模型

• 发表时间: 2022
• 作者: Gabriel Felipe Martinez

Science Dialogue

科学对话

• 发表时间: 2022
• 作者: Gabriel Felipe Martinez

iPSC Gene Editing as Applied to Regenerative Medicine

iPSC 基因编辑在再生医学中的应用

• 发表时间: 2022
• 作者: ウォルツェン ・クヌート

Considerations for gene editing in human iPS cells

人类 iPS 细胞基因编辑的注意事项

• 发表时间: 2022
• 作者: ウォルツェン ・クヌート

Precise genome editing in iPSCs

iPSC 中的精确基因组编辑

• 发表时间: 2022
• 作者: ウォルツェン ・クヌート