ChIP-seq experiments to define the role of MEIS transcription factors in cardiac differentiation
ChIP-seq experiments to define the role of MEIS transcription factors in cardiac differentiation
Cells were fixed with 1% HCHO for 10 minutes, neutralised with 0.125M Glycine for 5 min, washed twice with ice-cold PBS and spinned down. Cell pellets were snap frozen and stored at -80C. Embryonic stem (ES) cells were cultured on a monolayer of mitomycin inactivated mouse embryonic fibroblasts and maintained in DMEM/F12-based medium containing 20% knockout serum replacement (Invitrogen) and 10 ng/ml bFGF (Miltenyi Biotech). hPSCs were passaged using TrypLE Select (Invitrogen).  HESC NKX2-5eGFP/w and MEIS1/MEIS2 KO cell lines were differentiated using the embryoid body (EB) method as described in https://doi.org/10.1038/nmeth.1740. ChIPmentation samples and libraries were generated as described previously (https://www.doi.org/10.1016/j.xpro.2020.100187) with the use of the following antibodies: MEIS1 (Abcam ab19867; Active Motif MEIS1/2/3), GATA-6 (D61E4) XP Rabbit mAb (Cell Signaling Technology), H3K27ac (Abcam, ab4729) and rabbit polyclonal KMT2D (Atlas Antibodies). Purified DNA was amplified using Nextera indexes following manufacturer's protocol.