Wang Ruolei, Wu Yue, Zhou Ze, Ma Yicheng, Zhang Weidong, Wang Zihang, Luo Weihan, Hua Peng
State Key Laboratory of Reproductive Medicine and Offspring Health, Nanjing Medical University, Nanjing, China.
Front Cell Dev Biol. 2025 May 13;13:1572405. doi: 10.3389/fcell.2025.1572405. eCollection 2025.
Chromatin-protein interactions are fundamental for regulation of gene transcription. While chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) has long been the gold standard for mapping these interactions, emerging techniques such as CUT&RUN and CUT&Tag, which offer advantages such as low-input requirements and high signal-to-noise ratios, have aroused great attention. However, research addressing the potential biases introduced by enzyme-based tagmentation approaches and comparative assessment with ChIP-seq remain absent.
This study aims to systematically evaluate and compare the performance of ChIP-seq, CUT&Tag, and CUT&RUN for profiling genome-wide transcription factors and histone modification binding.
Our analysis revealed that all three methods reliably detect histone modifications and transcription factor enrichment, with CUT&Tag standing out for its comparatively higher signal-to-noise ratio. Detailed peak comparison revealed unique and overlapping enrichment among the three techniques. Additionally, CUT&Tag can identify novel CTCF peaks compared with the other two methods. A strong correlation was observed between CUT&Tag signal intensity and chromatin accessibility, highlighting its ability to generate high-resolution signals in accessible regions.
The systematic comparison summarizes the differences between CUT&Tag and CUT&RUN in terms of the signal-to-noise ratio and bias toward accessible chromatin. Considering the experimental procedures, signal specificity, and inherent biases, we recommend tailoring the choice of method to the type of chromatin-protein interaction under study. CUT&Tag offers a promising alternative for applications requiring high sensitivity and reduced background noise.
染色质与蛋白质的相互作用是基因转录调控的基础。长期以来，染色质免疫沉淀结合深度测序（ChIP-seq）一直是绘制这些相互作用图谱的金标准，而诸如CUT&RUN和CUT&Tag等新兴技术，因其具有低输入要求和高信噪比等优势，引起了广泛关注。然而，关于基于酶切标签法引入的潜在偏差以及与ChIP-seq的比较评估的研究仍然缺乏。
本研究旨在系统评估和比较ChIP-seq、CUT&Tag和CUT&RUN在全基因组范围内分析转录因子和组蛋白修饰结合情况的性能。
我们的分析表明，这三种方法都能可靠地检测到组蛋白修饰和转录因子富集，其中CUT&Tag的信噪比相对较高。详细的峰比较揭示了这三种技术之间独特的和重叠的富集情况。此外，与其他两种方法相比，CUT&Tag能够识别新的CTCF峰。在CUT&Tag信号强度与染色质可及性之间观察到强烈的相关性，突出了其在可及区域产生高分辨率信号的能力。
该系统比较总结了CUT&Tag和CUT&RUN在信噪比和对可及染色质的偏向性方面的差异。考虑到实验步骤、信号特异性和固有偏差，我们建议根据所研究的染色质-蛋白质相互作用的类型来选择合适的方法。对于需要高灵敏度和降低背景噪声的应用，CUT&Tag提供了一个有前景的替代方案。
