GSM5608498: siYAPsiTAZ_TEAD1_Rep1; Homo sapiens; OTHER
See Skene PJ et al 2018 (PMID: 29651053) 250,000 cells were bound to 10 µl concanavalin-A beads and collected into a 0.2 ml PCR tube. Antibody binding was conducted in the volume of 50 µl on thermomixer for 96-well PCR plates (Eppendorf) in 4 °C cold room for 2 hours, and Protein A-MNase binding for 1 hour. Wash buffer volume was adjusted to 150 µl. The digestion was performed under high Ca2+/low salt condition on ice for 15 minutes, followed by incubation at 37 °C for 30 min to release CUT&RUN fragments. Antibodies were diluted 1/100 . For histone marks profiling, CUT&RUN fragments were purified by spin column-based methods (Zymo DNA clean & concentrator-5 kit). For transcription factors and BAF complex profiling, CUT&RUN fragments were extracted by phenol-chloroform to recover small fragments. The libraries were generated by using the NEBNext ultraII DNA library prep kit for Illumina coupled with NEBNext multiplex oligos for Illumina (New England Biolabs) with few modifications optimized for small fragments (detailed in dx.doi.org/10.17504/protocols.io.wvgfe3w.)