TCC was performed as described.25 (link) Briefly, approximately 5e+07 PANC1 cells (ATCC, Manassas, VA, USA) were crosslinked with 1% formaldehyde for 10 minutes at room temperature, crosslinking was quenched with 0.125 M glycine for 5 minutes at room temperature, and cell pellets were collected and stored at –80°C. Nuclei were digested with 2000U HindIII (New England BioLabs, Ipswich, MA) before dilute solid-surface ligation reactions and TCC library preparation as described.25 (link) For drug treatments, PANC1 cells were grown to 60% confluency before a 24 hour treatment with 10 μM ICG-001 (Tocris, Bristol, UK), 10 μM C646 (Sigma-Aldrich, St. Louis, MO) or DMSO and fixed and harvested as described above. TCC libraries were sequenced using an Illumina HiSeq 2000.
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Chromatin Conformation Capture in PANC1 Cells
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Corresponding Organization : The University of Texas Health Science Center at San Antonio
Other organizations : University of Southern California, Oslo University Hospital, The Ohio State University
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Variable analysis
independent variables
- Drug treatments: 10 μM ICG-001, 10 μM C646, or DMSO
- Chromatin interactions measured by TCC
- PANC1 cells (ATCC, Manassas, VA, USA)
- Crosslinking with 1% formaldehyde for 10 minutes at room temperature
- Quenching of crosslinking with 0.125 M glycine for 5 minutes at room temperature
- Nuclei digestion with 2000U HindIII (New England BioLabs, Ipswich, MA)
- Dilute solid-surface ligation reactions and TCC library preparation
- PANC1 cells grown to 60% confluency before treatment
- Positive control: None specified
- Negative control: DMSO treatment
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