GSM9287349: Peripheral CD8+ T cells, siDUSP2 transfected, anti-CD3/CD28 activated, 24-hours_healthy donor 1; Homo sapiens; ChIP-Seq
1x106 nucleofected CD8+ T cells were fixed with 1% formaldehyde for 10 minutes, then quenched with glycine, and lysed in RIPA prior to sonication. Partially fragmented chromatin was then incubated with ETS1 (clone [D8O8A], Cell Signaling TECHNOLOGY) coupled Protein A and G Dynabeads (Invitrogen) overnight prior to magnetic bead separation and subjected to tagmentation (Illumina Tagment Kit) for 40 minutes at 37 degrees Celsius and overnight proteinase K (Thermo Scientific) treatment. The aqueous phase was then extracted by phenol:chloroform and precipitated with sodium acetation, glycogen, and ethanol incubated overnight prior to pelleting and resuspension in TE buffer. DNA was amplified with a High-Fidelity PCR Mix (NEW ENGLAND Bilabs) and 0.4 uM of primers for 12 cycles. Libraries were measured for concentration and quality by Qubit fluorometry and the Agilent Fragment Analyzer and normalized to 1.6 nM. Libraries were pooled and loaded onto a NovaSeq SP flow cell at a final concentration of 320 pM and the flow cell was sequenced as 50 x 2 paired end reads.