GSM9214453: HeLa cells, Uninfected, biol rep 3; Homo sapiens; ATAC-seq
At 72 hours post infection, uninfected and infected HeLa cells were washed twice with pre-warmed media, washed once with PBS, and detached via trypsinization. All samples had at least 93% cell viability based on trypan blue exclusion. 50,000 cells per sample were centrifuged at 1000 x g for 5 min at 4C, washed with 100 uL of cold PBS, and resuspended in 100 uL cold ATAC-seq lysis buffer (ActiveMotif). Lysed samples were immediately centrifuged at 1000 x g for 10 min at 4C and the pellets were resuspended in 50 uL tagmentation master mix per sample (2X tagmentation buffer [ActiveMotif], 10X PBS, 1% digitonin, 10% Tween-20, and 5 uL loaded Tn5). The tagmentation reactions were mixed by thorough pipetting and incubated at 37C with shaking at 400 rpm for 30 min. Reactions were purified with a MinElute PCR Purification Kit (Qiagen) and eluted in 10mM Tris buffer, pH 8. Tagmented DNA was sent to Novogene for library amplification and next-generation sequencing according to previously established protocols [PMID: 24097267, PMID: 28846090]. PCR amplification was performed with equimolar amounts of i5/i7 primers. PCR products were purified with AMPure beads and library quality was assessed with Qubit. Clustering of index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumina). After cluster generation, the library preparations were sequenced on an Illumina Hiseq platform and 150 bp paired-end reads were generated.