GSM7615607: IgG Control - FLAG-Dsup - CUT&RUN; Saccharomyces cerevisiae; OTHER
Nuclei from yeast cells expressing Dsup or mutant Dsup were purified according to published methods (doi:10.1007/978-1-0716-2257-5_9) with slight modifications described above. 1 mL aliquots containing approximately 5 x 107 nuclei were slow-frozen in an isopropanol chamber at  -80°C overnight. Immediately prior to an experiment, nuclei were thawed rapidly for 2-3 minutes at 37°C, then 100 μL of nuclei suspension was used per CUT&RUN reaction with the CUTANA™ ChIC/CUT&RUN Kit version 3 (EpiCypher). The protocol was carried out as directed by the version 3.2 user manual. MNase digestion of chromatin was performed for 2 hours at 4°C. See referenced text for further detail. 5 ng of DNA was used to prepare sequencing libraries with the Ultra II DNA Library Prep Kit (NEB #E7645L). Libraries were sequenced on an Illumina NextSeq 2000 platform. See referenced text for further detail.