title: GSM3316257: ISC-Ring1b_ChIPseq; Mus musculus; ChIP-Seq
library_construction_protocol: Intestinal epithelium from LGR5-Ring1a/b ++ mice was extracted following the protocol previously described. Single cells were resuspended in Sorting medium (as previously described) and LGR5-GFP positive cells were FACS-sorted with either FACS-Melody (‎BD FACSMelody Cell Sorter) or FACS-Jazz (‎BD FACSJAZZ Cell Sorter), discriminating only singlets (plotting Area vs Height on both SSC and FSC) and only alive cells with Propidium Iodide. An amount of 2.5 Milion cells were resuspended and sonicated with Branson Digital Sonifier (Branson) in IP buffer (SDS Buffer: Triton Buffer 2:1 – SDS Buffer (NaCl 100mM, Tris-HCl pH 8.1 50mM, NaN3 0.2%, SDS 0.5%); Triton Buffer ( NaCl 100mM, Tris-HCl pH 8.6 100mM, EDTA 5mM, NaN3 0.2%, Triton X-100 5%). Sonicated chromatin were incubated with 10ug of Ring1b antibody generated in the lab (Chiacchiera, Rossi, Sriganesh Jammula, et al. 2016) over night and the immunocomplexes were recovered by magnetic beads (Dynabeds, Life-technologies) and decrosslinked over night with NaHCO3 0.1M and 1% SDS. Decrosslinked DNA were purified.
attributes.tag: GEO Accession
attributes.value: GSM3316257