GSM4944233: Wild-type (Rubinstein-Taybi cohort) replicate 6; Mus musculus; ATAC-seq
Peripheral blood was obtained from facial vein bleed and CD19+ cells were isolated by positive selection using magnetic microbeads (Miltenyi 130-052-201) per manufacturer protocols and placed on ice. 5k cells were used per reaction, resuspended in 1xPBS and quickly spun to remove residual EDTA from isolation steps. Samples were resuspended in tagmentation reaction mix for 30 min (2.5uL TD1, 1X TD Buffer, Illumina Nextera DNA, FC-121-1030; .25uL 1% digitonin, Promega G9441; 1xPBS; ) gently shaking (300rpm on Eppendorf thermomixer) at 37'. We purified reactions using Zymo DNA Clean and Concentrator-5 kit (Zymo D4013) following manufacturer protocols and eluted with 10.5uL water to recover 10uL. Each reaction was then amplified and indexed per standard ATAC-seq protocol, total sample amplification cycles range from 6-10 cycles. After indexing and amplification, we purified samples using Select-A-Size purification columns (Zymo D4080) with a cutoff of 150bp to remove adapter dimers to allow for efficient sequencing on patterned flow cells, checked library size on BioAnalyzer using DNA High Sensitivity reagents (Agilent 5067-4626) and determined concentration using Qubit dsDNA HS Assay Kit (ThermoFisher Q32851).
GEO Accession
GSM4944233