title: GSM2679757: RAL_D225; Homo sapiens; ATAC-seq
library_construction_protocol: 50,000 sorted DCs from each condition were immediately prepped for ATAC-seq (Buenrostro et al., 2013).  Cells were pelleted by spinning at 500g for 5min at 4C.  Cells were washed in cold PBS and centrifuged again at 500g for 5min at 4C.  Cell pellets were lysed in cold lysis buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2 and 0.1% IGEPAL CA-630) and immediately spun at 500g for 10 min at 4C.  The nuclear pellet was used directly for the transposition reaction by resuspending in a reaction mix (20 μL 2× TD buffer, 2 μL Tn5 transposase (Illumina) and 18 μL nuclease-free water).  Transposition was performed for 30 min at 37C and sample DNA was purified using a MinElute kit (Qiagen). Library fragments were amplified by PCR using NEBnext PCR master mix (Illumina) and custom Nextera primers using the following conditions: 72 °C for 5 min; 98 °C for 30 s; and then cycling at 98 °C for 10 s, 63 °C for 30 s and 72 °C for 1 min. To stop amplification before saturation in order to reduce bias, we monitored the PCR reaction using SybrGreen-based qPCR.  Samples were cycled an average of 15 times.
attributes.tag: GEO Accession
attributes.value: GSM2679757