MRC Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK.
MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK.
Nat Commun. 2021 Jan 22;12(1):531. doi: 10.1038/s41467-020-20809-6.
Chromosome conformation capture (3C) provides an adaptable tool for studying diverse biological questions. Current 3C methods generally provide either low-resolution interaction profiles across the entire genome, or high-resolution interaction profiles at limited numbers of loci. Due to technical limitations, generation of reproducible high-resolution interaction profiles has not been achieved at genome-wide scale. Here, to overcome this barrier, we systematically test each step of 3C and report two improvements over current methods. We show that up to 30% of reporter events generated using the popular in situ 3C method arise from ligations between two individual nuclei, but this noise can be almost entirely eliminated by isolating intact nuclei after ligation. Using Nuclear-Titrated Capture-C, we generate reproducible high-resolution genome-wide 3C interaction profiles by targeting 8055 gene promoters in erythroid cells. By pairing high-resolution 3C interaction calls with nascent gene expression we interrogate the role of promoter hubs and super-enhancers in gene regulation.
染色质构象捕获（3C）为研究各种生物学问题提供了一种适应性强的工具。目前的 3C 方法通常提供整个基因组的低分辨率相互作用图谱，或者在有限数量的基因座上提供高分辨率相互作用图谱。由于技术限制，尚未在全基因组范围内实现可重复的高分辨率相互作用图谱的生成。在这里，为了克服这一障碍，我们系统地测试了 3C 的每一步，并报告了对当前方法的两项改进。我们表明，使用流行的原位 3C 方法生成的报告事件中，多达 30%是来自两个单个核之间的连接，但通过在连接后分离完整的核，可以几乎完全消除这种噪声。使用核滴定捕获 C，我们通过靶向红细胞中 8055 个基因启动子生成可重复的高分辨率全基因组 3C 相互作用图谱。通过将高分辨率 3C 相互作用调用与新生基因表达配对，我们研究了启动子枢纽和超级增强子在基因调控中的作用。
