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DiMeLo-cito:用于绘制蛋白质-DNA相互作用的单管协议揭示了CTCF在线粒分裂中的书签

Nathan Gamarra¹, Cy Chittenden¹, Kousik Sundararajan²

¹Stanford University, Department of Genetics, Palo Alto, California 94304, USA.

bioRxiv : the preprint server for biology

|March 31, 2025

概括

一种新的无洗方法,DiMeLo-cito,使用长读序列测序绘制蛋白质-DNA相互作用的地图. 这项技术揭示了CTCF与线粒染色体的结合,这对于跨细胞分裂的基因组稳定性至关重要.

科学领域:

基因组学就是基因组学.
分子生物学分子生物学
表观遗传学在表观遗传学中,表观遗传学是指表观遗传学.

背景情况:

基因组调节涉及复杂的蛋白质-DNA相互作用.
第三代测序为这些相互作用提供了强大的全基因组映射.
现有的方法,如用长读序列 (DiMeLo-seq) 进行定向甲基化,由于加工过程中的样本损失而存在局限性.

研究的目的:

开发一种新的,高效的协议,使用长读序列测序来绘制蛋白质-DNA相互作用.
克服以前方法的局限性,特别是样本损失.
为了实现高质量的全基因组蛋白质结合分析,从不同的细胞类型,包括在线粒分裂期间.

主要方法:

开发了DiMeLo-cito,这是一种单管,无洗的协议,用于长读测序.
应用DiMeLo-cito来检测基因组范围内的蛋白质结合,使用最小的细胞输入 (100,000 个细胞).
在不需要核外的情况下,分析线粒染色体中的蛋白质-DNA相互作用.

主要成果:

DiMeLo-cito最大限度地提高了基因组DNA产量和质量,以便长时间读取测序.
该协议成功检测出CTCF在双胞胎人体细胞中与线粒染色体的强结合.
这与卡略型不稳定的癌细胞系的发现形成鲜明对比,这表明CTCF在基因组架构维护中的作用.

结论:

DiMeLo-cito扩大了长读测序的适用性,用于研究基因组调节.
对线粒染色体的CTCF结合表明它在"书签"基因组稳定性关键部位中的作用.
这种方法为基因组组织和细胞分裂的调节提供了新的见解.

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