The Hi-C technique is widely employed to study the 3-dimensional chromatin architecture and to assemble genomes. The conventional in situ Hi-C protocol employs restriction enzymes to digest chromatin, which results in nonuniform genomic coverage. Using sequence-agnostic restriction enzymes, such as DNAse I, could help to overcome this limitation. In this study, we compare different DNAse Hi-C protocols and identify the critical steps that significantly affect the efficiency of the protocol. In particular, we show that the SDS quenching strategy strongly affects subsequent chromatin digestion. The presence of biotinylated oligonucleotide adapters may lead to ligase reaction by-products, which can be avoided by rational design of the adapter sequences. Moreover, the use of nucleotide-exchange enzymes for biotin fill-in enables simultaneous labelling and repair of DNA ends, similar to the conventional Hi-C protocol. These improvements simplify the protocol, making it less expensive and time-consuming. We propose a new robust protocol for the preparation of DNAse Hi-C libraries from cultured human cells and blood samples supplemented with experimental controls and computational tools for the evaluation of library quality.

中文翻译：

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DNase Hi-C 食谱

Hi-C 技术被广泛用于研究 3 维染色质结构和组装基因组。传统的原位 Hi-C 协议采用限制性内切酶消化染色质，导致基因组覆盖不均匀。使用与序列无关的限制性内切酶，例如 DNAse I，可以帮助克服这一限制。在本研究中，我们比较了不同的 DNAse Hi-C 协议，并确定了显着影响协议效率的关键步骤。特别是，我们表明 SDS 淬火策略强烈影响随后的染色质消化。生物素化寡核苷酸接头的存在可能导致连接酶反应副产物，这可以通过接头序列的合理设计来避免。而且，使用核苷酸交换酶进行生物素填充可以同时标记和修复 DNA 末端，类似于传统的 Hi-C 协议。这些改进简化了协议，降低了成本和耗时。我们提出了一种新的稳健协议，用于从培养的人类细胞和血液样本中制备 DNAse Hi-C 文库，并辅以用于评估文库质量的实验控制和计算工具。