GSM6935178: H3K4me3_CUT&RUN_KO; Mus musculus; OTHER
CUT&RUN was performed on 300,000 cells/sample using the ChIC/CUT&RUN Kit (EpiCypher #14-1048) and anti-H3K4me3 (EpiCypher Cat. No. 13-0041). The procedure followed the manufacturer's protocol with the following modifications. 1 μl anti-H3K4me3 antibody/sample was used. Following spike-in controls were used: K-MetStat Panel (2 μl of stock diluted 1:3) and 0.37 μl E. coli. pAG-MNase binding was performed for 20 min at RT while rotating. MNase digestion was performed for 120 min at 4C while rotating. Sequencing libraries were prepared with the NEBNext Ultra II Library Prep Kit (New England Biolabs Cat. No. E7645) and the Magnetic SPRI beads (Beckman Coulter Cat. No. B23317) for purification. PCR amplification conditions were: 14 cycles of DNA melting at 98˚C for 15 seconds, annealing plus extension at 60˚C for 10 seconds, followed by final extension at 72˚C for 1 minute. Libraries were quality-checked and quantified by Bioanalyzer (Agilent) and Qubit (ThermoFisher) and sequenced as PE150 on Illumina NovaSeq 6000.