GSM7025965: 4CBS_Droplet_Paired-Tag_H3K27me3_DNA; Mus musculus; OTHER
To prepare nuclei, mESC were dissociated with Accutase (Innovative Cell Technologies, AT104), collected by centrifugation, washed twice with PBS (Gibco, 10010023) then resuspended in cold nuclei permeabilization buffer (10 mM Tris-HCl pH 7.4 (Sigma, T4661), 10 mM NaCl (Sigma, S7653), 3 mM MgCl2 (Sigma, 63069), 1×Protease Inhibitor (Roche, 05056489001), 0.5 U µl−1 RNase OUT (Invitrogen, 10777-019), 0.5 U µl−1 SUPERase Inhibitor (Invitrogen, AM2694), 0.1% IGEPAL CA630 (Sigma, I8896) and 0.02% Digitonin (Sigma, Cat#D141)) for 1 minute. Nuclei were counted using a BioRad TC20 cell counter with 0.4% Trypan Blue (Gibco, 15250061) staining. For each experiment, around 0.5 million nuclei were aliquoted, and centrifuged for 10 min at 1,000g, 4 °C before proceeding to Droplet Paired-Tag experiments. Final DNA and RNA libraries amplification were performed according to the Chromium Single Cell ATAC Library kit manual, except that we used an increased number of amplification cycles (in total 12-13) for DNA (histone modality) library.