GSM6189880: ATAC_T8_IRF9KO_IFNb_01h30min_TDAL013; Mus musculus; ATAC-seq
Chromatin accessibility mapping by ATAC-seq was performed as previously described (Buenrostro et al., 2013; Corces et al., 2016), with minor adaptations. After centrifugation, the pellet was carefully resuspended in the transposase reaction mix (12.5µl 2xTD buffer, 2µl TDE1 (Illumina), 10.25µl nuclease-free water, and 0.125µL 10% NP-40 (Sigma) for macrophages and dendritic cells or 0.25µl 1% digitonin (Promega) for all other cell types) for 30 min at 37 °C. Following DNA purification with the MinElute kit eluting in 11µl, 1µl of the eluted DNA was used in a quantitative PCR reaction to estimate the optimum number of amplification cycles. The remaining 10μl of each library were amplified for the number of cycles corresponding to the Cq value from the qPCR (i.e., the cycle number at which fluorescence has increased above background levels, rounded down). Library amplification was followed by SPRI beads (Beckman Coulter) size selection to exclude fragments larger than 1,200bp. DNA concentration was measured with a Qubit fluorometer (Life Technologies). Library amplification was performed using custom Nextera primers (Buenrostro et al., 2013). The libraries were sequenced by the Biomedical Sequencing Facility at CeMM using the Illumina HiSeq 3000/4000 platform and the 50bp single-end configuration.