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| Status |
Public on Sep 23, 2020 |
| Title |
Differential H3K27ac ChIP-seq Analysis of HCT116 Cells In Response To Interferon Beta |
| Organism |
Homo sapiens |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
To show the utility of our method, MEIRLOP, we performed a differential motif enrichment analysis of regulatory elements modulated by interferon beta (IFN-β) treatment in HCT116 cells as measured by H3K27ac ChIP-seq.
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| Overall design |
Cell culture and treatment: HCT116 CMV-osTIR1 RAD21-mAC cells were obtained from Masato T. Kanemaki and cultured in McCoy’s 5A medium supplemented with 10% FBS. Cells were grown in a 37˚C incubator with 5% CO2. Cells were treated with 0.1% final DMSO for 6 hours and then treated with either IFN-β (1000 unit/ml) (n=2) for one hour or not further treated (n=2).
ChIP-seq and Crosslinking: Crosslinking and ChIP-seq were performed as described in Heinz et al., 2018 with few adjustments. Briefly, cells were fixed directly by adding formaldehyde into media to a final concentration of 1% formaldehyde for 10min at room temperature and quenched with 125mM Glycine. Cells were then pelleted at 300g for 5min at 4˚C, washed twice with cold PBS (with 0.5% BSA), snap frozen in liquid nitrogen and stored at -80˚C. ChIP-seq was performed on 500,000 cells as described in Heinz et al., 2018. H3K27ac antibodies were obtained from Active Motif (cat#:39133). Libraries were single-end sequenced for 84bp to a depth of 5.5 - 8.6 reads on an Illumina NextSeq500 instrument.
Differential ChIP-seq analysis: After sequencing and adapter trimming with FastP, 5.5M - 8.6M reads per library were aligned to the GRCh38 reference genome using bowtie2 at overall alignment rates of 98.5% - 99.5%. For each stimulation (n = 2) and control sample (n = 2), MACS2 was used to call peaks of median length 1kbp relative to a background input sample (of 16M reads). DiffBind was used to call differentially acetylated peaks between stimulated and unstimulated conditions.
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| Contributor(s) |
Delos Santos NP, Texari L, Benner C |
| Citation(s) |
32938397 |
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https://doi.org/10.1186/s12859-020-03739-4
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| Submission date |
Mar 29, 2020 |
| Last update date |
Oct 02, 2020 |
| Contact name |
Nathaniel Delos Santos |
| E-mail(s) |
nathaniel.p.delossantos@jacobs.ucsd.edu
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| Phone |
408-425-5535
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| Organization name |
University of California San Diego
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| Street address |
9500 Gilman Drive MC 0640
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| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92093 |
| Country |
USA |
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| Platforms (1) |
| GPL18573 |
Illumina NextSeq 500 (Homo sapiens) |
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| Samples (5)
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| GSM4443577 |
human_hct116_chip_h3k27ac-dmso-ifnb-60min-r1_LT |
| GSM4443578 |
human_hct116_chip_h3k27ac-dmso-ifnb-60min-r2_LT |
| GSM4443579 |
human_hct116_chip_h3k27ac-dmso-mock-0min-r1_LT |
| GSM4443580 |
human_hct116_chip_h3k27ac-dmso-mock-0min-r2_LT |
| GSM4443581 |
human_hct116_chip_input-dmso-mock-60min-r1_LT |
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| Relations |
| BioProject |
PRJNA616157 |
| SRA |
SRP254492 |
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