Ultrasensitive multiomic platform using epitope-targeted DNA methylation mapping

使用表位靶向 DNA 甲基化作图的超灵敏多组学平台

基本信息

  • 批准号:
    10758061
  • 负责人:
  • 金额:
    $ 103.57万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2022
  • 资助国家:
    美国
  • 起止时间:
    2022-02-01 至 2025-01-31
  • 项目状态:
    未结题

项目摘要

PROJECT SUMMARY Gene expression is regulated by the complex molecular cross-talk between DNA methylation (DNAme) and other chromatin features: e.g. histone post-translational modifications (PTMs) and chromatin associated proteins (ChAPs; transcription factors and chromatin remodelers). Significantly, changes in the chromatin landscape can have a profound impact on DNAme patterning (and vice versa), and these changes are connected to development as well as a broad range of diseases (from cancer to neurological disorders). However, our understanding of how DNAme co-occurs / coordinates with additional chromatin features to control gene expression is limited by a lack of reliable genomic tools. Here, EpiCypher is partnering with New England Biolabs (NEB) to develop Targeted Enzymatic Methylation-sequencing (TEM-seqTM), an ultra-sensitive multiomic mapping technology that delivers high resolution DNAme profiles (5mC/5hmC) at epitope-defined chromatin features. EpiCypher is leading the development of ultra-sensitive genomic mapping assays that use CUT&RUN / CUT&Tag methods (under the CUTANA® platform) to generate truly quantitative data using dramatically reduced cell input and sequencing depth (>10-fold savings on each parameter vs. ChIP-seq). CUTANA assays are supported by EpiCypher’s proprietary spike-in designer nucleosome (dNuc) technology to enable technical monitoring and quantitative normalization. The key innovation of the TEM-seq project is the development of a novel multiomic workflow that marries EpiCypher’s quantitative CUTANA CUT&RUN technology with unbiased DNAme analysis using NEB’s enzymatic methyl-seq (EM-seq) approach. EM-seq utilizes the enzymatic conversion of DNAme (5mC / 5hmC) and provides a much-needed alternative to bisulfite sequencing (BS; a chemical treatment that degrades DNA and has systemic sequence biases) to generate high resolution, unbiased DNAme profiles with ~10-fold less sample input (vs. BS). In Phase I Aim 1, we will rigorously validate our TEM-seq workflow in three cell lines, benchmark results against standard CUT&RUN and EM-seq assays, and further develop EpiCypher’s spike-in controls for compatibility with TEM-seq. We will advance to Phase II when we demonstrate that TEM-seq generates highly reliable DNAme maps associated with histone PTMs and ChAPs using <50k cells and <10M reads. In Phase II Aim 2, we will expand development of spike-in control panels and develop robust protocols for a wide panel of chromatin features (using validated antibodies) and sample processing methods (fresh, frozen, and fixed), including drug treatment time-course experiments to enable clinical applications. In Phase II Aim 3, we will develop / validate a TEM-seq beta kit, and also create a data analysis portal and automated assays to accelerate commercial adoption and enable a high-throughput service offering. TEM-seq will provide a powerful new tool to expand our understanding of complex chromatin signaling, further unlocking the potential of epigenetics-targeted drugs and diagnostics.
项目摘要 基因表达受DNA甲基化(DNAme)之间复杂的分子串扰调节。 和其他染色质特征:例如组蛋白翻译后修饰(PTM)和染色质相关的 蛋白质(ChAP;转录因子和染色质重塑)。值得注意的是, 景观可以对DNA模式产生深远的影响(反之亦然),这些变化是相互关联的。 发展以及广泛的疾病(从癌症到神经系统疾病)。但我们的 理解DNAme如何与其他染色质特征共同发生/协调以控制基因 由于缺乏可靠的基因组工具,表达受到限制。在这里,EpiCypher与新英格兰生物实验室合作 (NEB)开发靶向酶促甲基化测序(TEM-seqTM),一种超灵敏的多组 在表位定义的位置提供高分辨率DNAme图谱(5 mC/5 hmC)的映射技术 染色质特征。EpiCypher正在领导超灵敏基因组图谱分析的开发, CUT&RUN / CUT&Tag方法(在CUTANA®平台下),使用 显著降低细胞输入和测序深度(与ChIP-seq相比,每个参数节省>10倍)。 CUTANA检测得到EpiCypher专有的加标设计核小体(dNuc)技术的支持, 实现技术监控和量化标准化。TEM-seq项目的关键创新在于 开发一种新型的多组学工作流程,结合EpiCypher的定量CUTANA CUT&RUN 使用NEB的酶促甲基测序(EM-seq)方法进行无偏DNA分析的技术。EM-seq 利用DNAme的酶促转化(5 mC/5 hmC),并提供急需的亚硫酸氢盐替代品 测序(BS;一种降解DNA并具有系统性序列偏差的化学处理),以产生高水平的 高分辨率、无偏的DNAme图谱,样品输入量约为BS的1/10。在第一阶段目标1中,我们将严格 在三种细胞系中验证我们的TEM-seq工作流程,将结果与标准CUT&RUN和EM-seq进行基准测试 分析,并进一步开发EpiCypher的spike-in控制与TEM-seq兼容。我们将推进到 第二阶段,当我们证明TEM-seq生成与组蛋白相关的高度可靠的DNAme图谱时, 使用<50 k细胞和<10 M读数的PTM和ChAP。在第二阶段目标2中,我们将扩大 控制面板,并为广泛的染色质特征面板开发强大的协议(使用经验证的抗体) 和样品处理方法(新鲜、冷冻和固定),包括药物处理时程实验, 实现临床应用。在第二阶段目标3中,我们将开发/验证TEM-seq beta试剂盒,并创建一个 数据分析门户和自动化分析,以加速商业应用并实现高通量 服务提供。TEM-seq将提供一个强大的新工具来扩展我们对复杂染色质的理解 信号,进一步释放表观遗传学靶向药物和诊断的潜力。

项目成果

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Michael-Christopher Keogh其他文献

Michael-Christopher Keogh的其他文献

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{{ truncateString('Michael-Christopher Keogh', 18)}}的其他基金

Scalable and quantitative chromatin profiling from formalin-fixed paraffin-embedded samples
对福尔马林固定石蜡包埋样品进行可扩展和定量的染色质分析
  • 批准号:
    10696343
  • 财政年份:
    2023
  • 资助金额:
    $ 103.57万
  • 项目类别:
Ultrasensitive multiomic platform using epitope-targeted DNA methylation mapping
使用表位靶向 DNA 甲基化作图的超灵敏多组学平台
  • 批准号:
    10833236
  • 财政年份:
    2023
  • 资助金额:
    $ 103.57万
  • 项目类别:
High-resolution genomic mapping of ssDNA and associated proteins for Alzheimer's disease research
用于阿尔茨海默病研究的 ssDNA 和相关蛋白的高分辨率基因组图谱
  • 批准号:
    10382044
  • 财政年份:
    2022
  • 资助金额:
    $ 103.57万
  • 项目类别:
Quantitative mapping of dynamic epigenetic states in rare and stimulated immune cells
稀有和刺激免疫细胞动态表观遗传状态的定量图谱
  • 批准号:
    10481225
  • 财政年份:
    2022
  • 资助金额:
    $ 103.57万
  • 项目类别:
Quantitative mapping of dynamic epigenetic states in rare and stimulated immune cells
稀有和刺激免疫细胞动态表观遗传状态的定量图谱
  • 批准号:
    10686135
  • 财政年份:
    2022
  • 资助金额:
    $ 103.57万
  • 项目类别:
Ultrasensitive multiomic platform using epitope-targeted DNA methylation mapping
使用表位靶向 DNA 甲基化作图的超灵敏多组学平台
  • 批准号:
    10384022
  • 财政年份:
    2022
  • 资助金额:
    $ 103.57万
  • 项目类别:
Ultrasensitive multiomic platform using epitope-targeted DNA methylation mapping
使用表位靶向 DNA 甲基化作图的超灵敏多组学平台
  • 批准号:
    10622310
  • 财政年份:
    2022
  • 资助金额:
    $ 103.57万
  • 项目类别:
A new epigenetic toolbox for inflammation research and drug discovery
用于炎症研究和药物发现的新表观遗传学工具箱
  • 批准号:
    10610898
  • 财政年份:
    2021
  • 资助金额:
    $ 103.57万
  • 项目类别:
A new epigenetic toolbox for inflammation research and drug discovery
用于炎症研究和药物发现的新表观遗传学工具箱
  • 批准号:
    10401943
  • 财政年份:
    2021
  • 资助金额:
    $ 103.57万
  • 项目类别:
A new epigenetic toolbox for inflammation research and drug discovery
用于炎症研究和药物发现的新表观遗传学工具箱
  • 批准号:
    10257054
  • 财政年份:
    2021
  • 资助金额:
    $ 103.57万
  • 项目类别:

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使用表位靶向 DNA 甲基化作图的超灵敏多组学平台
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    $ 103.57万
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