The functional implications of the three-dimensional genome organization are becoming increasingly recognized. The Hi-C and HiChIP research approaches belong among the most popular choices for probing long-range chromatin interactions. A few methodical protocols have been published so far, yet their reproducibility and efficiency may vary. Most importantly, the high frequency of the dangling ends may dramatically affect the number of usable reads mapped to valid interaction pairs. Additionally, more obstacles arise from the chromatin compactness of certain investigated cell types, such as primary T cells, which due to their small and compact nuclei, impede limitations for their use in various genomic approaches. Here we systematically optimized all the major steps of the HiChIP protocol in T cells. As a result, we reduced the number of dangling ends to nearly zero and increased the proportion of long-range interaction pairs. Moreover, using three different mouse genotypes and multiple biological replicates, we demonstrated the high reproducibility of the optimized protocol. Although our primary goal was to optimize HiChIP, we also successfully applied the optimized steps to Hi-C, given their significant protocol overlap. Overall, we describe the rationale behind every optimization step, followed by a detailed protocol for both HiChIP and Hi-C experiments.
SARS‐CoV‐2 Seroconversion in Humans: A Detailed Protocol for a Serological Assay, Antigen Production, and Test Setup
