GSM3610520: ChIP-seq_GRKO DexLPS rep2; Mus musculus; ChIP-Seq
ChIP was performed as previously described (Uhlenhaut et al., 2013) with the addition of a chromatin pre-clearing step. Briefly, cells were fixed with 30min DSG and 10min 1% formaldehyde. Crosslink was quenced with 0.2M glycine for 5min. Cells were lysed with Fast IP buffer (150mM NaCl, 5mM EDTA, 5mM Tris, 1% Triton X-100 and 0.5% NP40). Chromatin was sheared in shearing buffer (1% SDS, 10µM EDTA and 0.05% Tris) until majority of chromatin fragments were between 200-500bp. Chromatin pre-clearing was done by incubation with Dynabeads at 4°C rotating before IP set-up overnight. Next day, Dynabeads were added to IPs and 6h incubated at 4°C rotating. IPs were washed 6 times with Fast IP buffer. DNA was eluted with ChIP elution buffer (100mM NaHCO3 and 1%SDS). Library preperation was performed from 2 ng of ChIP DNA using the Kapa Hyper Prep Kit with PCR library amplification (#KK8504, KapaBiosystems) according to the manufactur's protocol. Shortly, ChIP DNA was end-repaired and A-tailed before adapter ligation. After adapter ligation, the ChIP DNA was purified with Agencourt AMPure XP beads (#A63880, Beckman Coulter) and size-selected for 360-600 bp using the Pippin Prep System from Saga Science. The size-selected library is amplified by PCR using the Kapa Hyper Prep Kit (#KK8504,KapaBiosystems) and purified terminally with Agencourt AMPure XP beads (#A63880, Beckman Coulter).
GEO
GSM3610520
GEO Accession
GSM3610520