Engineering locus-specific binders to DNA modifications

工程化位点特异性结合剂以进行 DNA 修饰

• 批准号: 10593668
• 负责人: Albert Keung
• 金额: $ 22.8万
• 依托单位: NORTH CAROLINA STATE UNIVERSITY RALEIGH
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10593668
• 关键词: Address; Affinity; Automobile Driving; Awareness; BCAR1 gene; Base Pairing; Binding; Binding Proteins; Biochemical; Biological; Biological Assay; Biotechnology; Cell physiology; Cells; Chemicals; Clustered Regularly Interspaced Short Palindromic Repeats; DNA; DNA Binding; DNA Modification Process; DNA Restriction-Modification Enzymes; Detection; Development; Diagnostic; Dimensions; Directed Molecular Evolution; Discipline; Disease; Engineering; Epigenetic Process; Etiology; Eukaryota; Event; Exhibits; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Engineering; Genome; Genomic DNA; Genomics; Hour; Immunologics; Immunoprecipitation; Life; Location; Major Groove; Maps; Measurement; Measures; Methylation; Modification; Molecular; Mutation; Nature; Nucleic Acids; Nucleotides; Pattern; Phenotype; Prokaryotic Cells; Protein Engineering; Proteins; RNA Splicing; Reagent; Reporting; Role; Safety; Sensitivity and Specificity; Site; Specificity; Stimulus; Structure; Surface; System; Technology; Therapeutic; Tissues; Work; Yeasts; Zinc Fingers; biological research; biological systems; bisulfite sequencing; cell type; epigenome; epigenome editing; experimental study; genome editing; genomic locus; imprint; innovation; insight; nanopore; novel; tool; whole genome; yeast two hybrid system

Project Summary Chemical modifications to genomic DNA are ubiquitous across all kingdoms of life. They are diverse in nature including methylation, hydroxymethylation, and formylation of nucleic acid residues. It is also increasingly clear that these modifications have diverse regulatory roles. In eukaryotes, DNA modifications regulate gene expression, RNA splicing, genome organization, and gene imprinting; these molecular functions also have important downstream consequences for development, disease, and organismal interactions with environmental stimuli and chemicals. Furthermore, with the advent of genome and epigenome editing technologies, these modifications can bias the specificity and efficiency of molecular tools. Given the broad importance of DNA modifications, any experimental tools that are able to control, measure, sense, or track them would have profound impacts across biological disciplines. What is particularly needed, but not yet technologically possibly, is the ability to simultaneously sense epigenetic modifications at specific genomic locations in living and single cells. This capability would unlock a broad palette of new experimental approaches to reveal new insights into topics such as gene imprinting, causation vs. correlation of epigenetic modifications, and cell and tissue stochasticity. Furthermore, it would inform the development of new classes of affinity reagents for biochemical assays, disease detection, and diagnostics and of genome and epigenome editing technologies that are aware of and specific towards DNA modification state. To drive this new realm of studies, we will create the first molecular tool that binds DNA hydroxymethylation at specific and programmable genomic loci, and provide a broadly applicable platform for engineering many other similar molecular reagents.

项目摘要 对基因组DNA的化学修饰在所有生命王国中都普遍存在。它们在性质上是多种多样的 包括核酸残基的甲基化、羟甲基化和甲基化。人们也越来越清楚， 这些修饰具有不同的调节作用。在真核生物中，DNA修饰调节基因表达， RNA剪接、基因组组织和基因印记;这些分子功能也具有重要的 对发育、疾病和生物体与环境刺激的相互作用的下游后果， 化学品此外，随着基因组和表观基因组编辑技术的出现，这些修饰可以 偏向分子工具的特异性和效率。鉴于DNA修饰的广泛重要性，任何 能够控制、测量、感知或跟踪它们的实验工具将对整个人类产生深远的影响。 生物学科。特别需要的，但在技术上还不可能的是， 同时感测活细胞和单细胞中特定基因组位置处的表观遗传修饰。这 这种能力将开启一个新的实验方法的广泛调色板，以揭示对主题的新见解， 基因印记、表观遗传修饰的因果关系与相关性以及细胞和组织的随机性。 此外，它将为开发用于生化测定、疾病诊断和治疗的新型亲和试剂提供信息。 检测和诊断以及基因组和表观基因组编辑技术， 走向DNA修饰状态。为了推动这一新的研究领域，我们将创造第一个分子工具， 在特定的和可编程的基因组位点结合DNA羟甲基化，并提供了广泛适用的 平台工程许多其他类似的分子试剂。