title: GSM3593231: ATACseq_HCAEC_24h_r2; Homo sapiens; ATAC-seq
library_construction_protocol: 5x10^4 cells were spun down at 500g for 5 minutes at 4°C. Whole cell pellets were subjected to a first round of cell membrane lysis using 50 μL of ice-cold hypotonic buffer (0.1% Sodium citrate tribasic dehydrate; 0.1% Triton X-100) and incubating on ice for 30 minutes. The hypotonic buffer was removed by centrifugation at 500g for 5 minutes at 4°C and we subsequently discarded the supernatant. Crude nuclei lysates were prepared by resuspending cells in lysis buffer (10mM Tris-HCl pH 7.4; 10mM NaCl; 3mM MgCl2; 0.1% Igepal CA-630 and incubating for 30 minutes on ice. Following the removal of lysis buffer by centrifugation at 500g for 5 minutes at °4C, transposition reaction of open chromatin was achieved by resuspending free nuclei in tagmentation mix (22.5 μL Tagment DNA Buffer; 2.5 μL Tagment DNA enzyme; 25 μL H2O) (Illumina) and incubating at 37°C for 30 minutes. Purification of DNA was performed with MinElute (Qiagen) according to the manufacturer's protocol. Barcoding and amplification was prepared using Nextera Index Kit (Illumina, FC-121-1011) with the following thermal profile: 30 seconds at 98°C and a three-step cycle of 10 seconds at 98°C, 30 seconds at 63°C and 1 minute at 72°C repeated 12 times followed by 5 minutes at 72°C. Amplified ATACseq libraries were purified using GeneRead Size Selection Kit (Qiagen).
attributes.tag: GEO Accession
attributes.value: GSM3593231