Rapid screening of gene-edited cells

快速筛选基因编辑细胞

• 批准号: 9751906
• 负责人: Bo Huang
• 金额: $ 20.06万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2020-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9751906
• 关键词: Address; Benchmarking; Biomedical Research; Cell Line; Cell Separation; Cells; ChIP-seq; Communities; Complement; Complex; DNA; DNA Integration; DNA Sequence; Detection; Economics; Elements; Engineering; Enzymes; Event; Fluorescence; Gene Expression; Genes; Genetic; Genome; Genome engineering; Genomic DNA; Genomics; Genotype; Goals; Knock-in; Label; Libraries; Mammalian Cell; Mediating; Methods; Pathway interactions; Procedures; Process; Proteins; Reporter; Reporting; Resources; Signal Transduction; System; TEV protease; Techniques; Technology; Time; Untranslated RNA; base; biological research; complement system; cost effective; genome-wide; imaging approach; rapid technique; repaired; scale up; screening; single molecule; site-specific integration

Project Summary Recently breakthroughs in gene editing technology have revolutionized many fields of biological and biomedical research, enabling applications including large-scale tagging of endogenous genes or editing of non-coding genomic elements. Screening of edited cell lines for those containing the correct edits, however, have mostly relied on the classical clonal selection method, which is slow and resource-intensive. Here, we propose to develop an enzymatic amplification technique inside living cells to report 1) the expression of a low abundance protein and 2) a single copy of specific DNA sequence in the genome. This technique will enable rapid isolation of edited cells by simple fluorescence-based cell sorting, thus greatly enhancing the efficiency and accessibility of gene editing for cell lines.

项目概要 最近基因编辑技术的突破彻底改变了生物和生物医学的许多领域 研究，支持包括大规模内源基因标记或非编码编辑在内的应用 基因组元件。然而，筛选编辑后的细胞系以寻找包含正确编辑的细胞系，主要是 依赖于经典的克隆选择方法，该方法速度慢且资源密集。在此，我们建议 开发活细胞内的酶扩增技术来报告 1) 低丰度的表达 蛋白质和 2) 基因组中特定 DNA 序列的单个副本。该技术将实现快速隔离 通过简单的基于荧光的细胞分选来编辑细胞，从而大大提高效率和可及性 细胞系的基因编辑。