ChIP-Atlas: SRX7626336

Antigen

Cell type Class: Others
Cell type: Mesenchymal Stromal Cells
MeSH Description: Mesenchymal stromal cells found in WHARTON JELLY, in the perivascular, intervascular, and subamnion regions of the UMBILICAL CORD. They are fibroblastic in appearance, rich in hyaluronic acid, and have been induced to form bone, cartilage, and adipose cells in vitro.

library_strategy: ChIP-Seq
library_source: GENOMIC
library_selection: ChIP
library_construction_protocol: Chromatin Immunoprecipitation of CTCF was performed using the ChIP-IT PBMC kit with the fixation protocol of the ChIP-IT High Sensitivity kit (both Active Motife, Carlsbad, CA, USA). In brief, MSCs of early passage (P 2, n = 3) and late passage (P 8 – 14, n = 3) were cross-linked with 1.2 % formaldehyde (Applichem, Darmstadt, Germany) for 15 min at room temperature. After cell lysis, chromatin fragmentation was performed by sonication for 20 min (20 cycles a´ 30 sec on/30 sec off, total on-time 10 min) in a water bath sonicator (Biorupter® Pico Sonicator Device, Diagenode). 5 - 33 µg of sonicated chromatin was incubated with 4 µg ChIP-validated CTCF antibody (Active Motif) overnight on an end-to-end rotator at 4°C. For immunoprecipitation, samples were incubated with protein G agarose beads (Active Motife) at 4°C for 3 hours. Finally, the cross-link was reversed, the DNA was proteinase K digested and purified. Sequencing and library preparation were conducted by the IZKF Sequencing Core Facility of RWTH Aachen University Medical school.