GSM6753086
For the preimplantation embryos: At the time of collection, the zona pellucida was remove using Tyrode's acid (Sigma, T1788-100ML), embryos were washed in M2 medium (Sigma, MR-015-D) and dissociated into single cells by incubating in TryplE (Thermo Fisher, 12605010) for approximately 5 minutes and pipetting up an down with a mouth pipet. Embryos were dissociated one by one and the resulting single cells were placed in M2 medium. They were then transferred to a 384-well plate (one single cell per well) containing 5uL of mineral oil and 100nL of CELseq primer (scDam&T) or 100nL of DamID buffer (0.15% NP40 and 2mM dNTPs) using a mouth pipet. For the mESC: Cells were trypsinized and harvested in medium with 20 μg/mL Hoechst 34580 (Sigma 63493) per 1x10^6 cells for 45 minutes at 37°C. The cells were stained for live/dead gating with 1ug/ml propidium iodide and singlets were index sorted based on their Hoechst profile. One cell per well was sorted into 384-well plates (Biorad, HSP3801) using the BD FACS Jazz cell sorter. Wells contained 5 μL filtered mineral oil (Sigma) and 50nL of 0.5uM unique CELseq primer. The scDam&T protocol and library preparation was performed as previously described in detail (PMID: 32350457).