GSM4058436: Nsd1KO_Nsd1-wt_Differentiation_H3K36me3_rep1; Mus musculus; ChIP-Seq
Pull-down beads were Protein G sepharose (Sigma-Aldrich, P3296). Washes were done with the low salt, high salt, li-Cl wash buffers according to EZ-Magna CHIP A/G kit protocol from Millipore. DNA elution was done with the IPURE Kit V2 (Diagenode, Cat.No. C03010012). Antibodies: GATA1 ab11862 10 ug per ChIP Abcam; H3K36me3: ab9050-11 10 ug per ChIP Abcam; H3K27ac: ab4729 10 ug per ChIP Abcam ChIP protocol was adapted from the EZ-Magna ChIPTM A/G kit protocol (Millipore, Merck KGaA, Darmstadt, Germany). Cells (Nsd1 WT and Nsd1 Set in maintenance as well as 24h differentiation) were fixed with 1% formaldehyde, lysed with Cell Lysis and then Nuclear Lysis buffers to a concentration of 20^106 cells per mL, and finally sonicated (20-min cycle on Covaris apparatus; KBioscience). Sheared chromatin was immunoprecipitated overnight using the following antibodies: anti-GATA1 (Abcam, ab11852), anti-H3K36me3 (Abcam, ab9050-100), anti-H3K27ac (Abcam, ab4729). 1/10 of the sheared chromatin was used as a reference (Input). Immune complex collection was performed using Protein G Sepharose (Sigma-Aldrich, P3296) for 1h30 at 4°C. Rinses were done according to Magna ChIPTM kit protocol with Low salt, High salt and LiCL immune complex wash buffers. Finally, elution was performed according the IPure Kit protocol (Diagenode, Cat.No. C03010012) following manufacturer's instructions. Two independent ChIP-seq experiments were conducted for Gata1, H3K27ac and H3K36me3.
GEO Accession
GSM4058436