title: GSM3731704: AKI1_CD8_Day-2_Rep2; Homo sapiens; ATAC-seq
library_construction_protocol: ATAC-seq library were prepared following the Omni-ATAC protocol. Briefly, frozen cells were thawed and stained for live cells using Ghost-Dye 710 (Tonbo Biosciences, CA, USA). 50,000 lived cells were FACS sorted and washed once with cold PBS. Technical replicates were done for most of the samples. Cell pellets were resuspended in 50µl cold ATAC-Resuspension buffer (10mM Tris-HCl (Sigmal Aldrich, MO, USA) pH 7.4, 10mM NaCl, 3mM MgCl2 (Sigma Aldrich,) containing 0.1% NP40 (Life Technologies, Carlsbad, CA), 0.1% Tween-20 (Sigma Aldrich) and 0.01% Digitonin (Promega, WI, USA) for 3 mins. Samples were washed once in cold resuspension buffer with 0.1% Tween 20, and centrifuged for 4C for 10 min. Extracted nuclei were resuspended in 50µl of Tn5 reaction buffer (1x TD buffer (Illumina, CA, USA), 100nM Tn5 Transposase (Ilumina), 0.01% Digitonin, 0.1% Tween-20, PBS and H20), and incubated at 37C for 30 min at 300rpm. Transposed samples were purified using MinElute PCR purification columns (Qiagen, Germany) as per manufacturer's protocol. Purified samples were amplified and indexed using custom Nextera barcoded PCR primers as described in (Buenrostro JD et al., Transposition of native chromatin for fast and sensitive epigenomic profile of open chromatin, DNA-binding proteins ad nucleosome position. Nature Methods, 2013). DNA libraries were purified using MinElute columns and pooled at equal molarity. To remove primer dimers, pooled libraries were further cleaned up using AmPure beads (Beckman Coulter, CA, USA).
attributes.tag: GEO Accession
attributes.value: GSM3731704