Cis-regulatory elements (cREs) and their long-range interactions are crucial for spatial-temporal gene regulation. While cREs can be characterized as accessible chromatin sequences, comprehensively identifying their spatial interactions remains a challenge. We recently developed a method, HiCAR (Hi-C on Accessible Regulatory DNA), which combines Tn5 transposase and chromatin proximity ligation to analyze open chromatin-anchored interactions in low-input cells. Application of HiCAR in human embryonic stem cells and lymphoblastoid cells reveals high-resolution chromatin contacts with efficiency comparable to in situ Hi-C across various distance ranges. Moreover, HiCAR was successfully applied to 30,000 primary human muscle stem cells, showcasing its potential for analyzing chromatin accessibility and looping in low-input primary cells and clinical samples. Here, we provide a detailed step-by-step protocol to perform the updated HiCAR experiments. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Tn5 Transposase Assembly Basic Protocol 2: HiCAR Library Preparation.
Keywords: Tn5; chromatin accessibility; chromatin interaction; chromosome conformation capture; restriction enzyme.
© 2023 Wiley Periodicals LLC.