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自动化样本准备用于单细胞基因组后翻译修饰 (sc-hPTM2) 的多重分析.

Giulia Barotti1, Ronald Cutler1, Joshua Cantlon2

  • 1Albert Einstein College of Medicine.

Journal of visualized experiments : JoVE
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概括

本研究引入了一种自动化的单细胞蛋白质组学工作流程,用于分析基因素翻译后修饰 (hPTMs). 该方法可以在单细胞水平上对染色质异质性和表观遗传变化进行敏感,可重复的研究.

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科学领域:

  • 表观遗传学和分子生物学
  • 蛋白质组学是指蛋白质组学.
  • 细胞生物学 细胞生物学

背景情况:

  • 基因组转化后修饰 (hPTMs) 调节染色体组织和基因表达.
  • 失调的HPTM与癌症,衰老和发育障碍有关.
  • 目前HPTM的蛋白质组学方法通常是批量分析,缺乏单细胞分辨率.

研究的目的:

  • 开发一个自动化,敏感的工作流程,用于单细胞质子PTM分析.
  • 为了克服低基因素丰度和单细胞中高修饰水平的技术挑战.
  • 为了能够在单细胞分辨率下调查染色质异质性和表观遗传扰乱.

主要方法:

  • 一个自动化纳米液体处理系统,用于纳米升级处理单个细胞.
  • 使用ArgC Ultra蛋白酶进行消化,避免素阻断衍生.
  • 一个双复合的标签策略,使用酸和其化模拟物进行定量比较.

主要成果:

  • 建立了一个简化和敏感的协议,用于单细胞质子PTM分析.
  • 工作流显示出高可重复性和最小的样本处理.
  • 该方法成功应用于分析酸盐治疗对肝细胞癌球状瘤的反应中的基因组乙化变化.

结论:

  • 开发的工作流提供了一个强大的工具,用于对基因素PTMs进行高分辨率分析.
  • 这一进步有助于研究细胞群内的表观遗传异质性.
  • 该协议为了解HPTM在各种生物过程和疾病中的作用开辟了新的途径.