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| Status |
Public on Nov 19, 2022 |
| Title |
Rep A-IP-siNT |
| Sample type |
SRA |
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| Source name |
HCT116 with ORF59 reporter
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| Organism |
Homo sapiens |
| Characteristics |
cell line: HCT116 colorectal line derivative
treatment: nontargeting siRNA, siNT
chip antibody: anti-RPB3 (Rabbit polyclonal, EMD Millipore, Cat#: ABE999, Lot #3764357)
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| Treatment protocol |
Cells were treated with 30nM of indicated siRNA 96 hrs prior to harvesting.
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| Growth protocol |
All cell lines were grown at 37°C with 5% CO2, with 1X penicillin-streptomycin (Sigma), and 2mM L-glutamine (Fisher) in DMEM (Sigma) supplemented with 10% FBS (Sigma);
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were fixed with 0.5% formaldehyde at RT for 10 min then quenched with 150 mM glycine at room temperature, 5 min. Cells were harvested by scraping in cold 1X PBS and pelleted at 1000 x g, 5 min at 4°C. Harvested cells were lysed for 30 min at 4°C in Farnham lysis buffer (5 mM PIPES, 85 mM KCl, 0.5% NP-40, 1 mM PMSF, 1X protease inhibitor cocktail (PIC)). Additional details of Dna shearing, and IP are given in Devlin et al. 2022.
For ChIP-seq library preparations, 4 ng of ChIP or input DNA was processed using the KAPA Hyper Prep Kit according to the manufacturer’s protocol (Roche KK8504).
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Single end sequenced reads were trimmed for adaptor sequence and filtered for quality with trim galore/0.6.4, and aligned to synthetic genome assembly (ORF59_reporter integatred into GRCh38 and dm6) using Bowtie2/2.3.2
The aligned reads were subsequently filtered for mapping quality (MAPQ 30) and PCR duplicates by SAMtools/1.6 and Picard. Uniquely mappable reads were retained for further analysis.
SpikeIN normalizations were performed after calculating scaling factor for each data set.
MACS2 was used for peak calling and generation of Input normalized (subtracted) coverage tracks.
Assembly: chimera genome assembly (ORF59_reporter integrated into chr19:55,115,766-55,121,825 of GRCh38)
Supplementary files format and content: Normalized Bigwig files
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| Submission date |
Apr 18, 2022 |
| Last update date |
Nov 19, 2022 |
| Contact name |
Nicholas K Conrad |
| E-mail(s) |
nicholas.conrad@utsouthwestern.edu
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| Phone |
2146480795
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| Organization name |
UT Southwestern Medical Center
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| Department |
Microbiology
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| Street address |
6000 Harry Hines Blvd
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| City |
Dallas |
| State/province |
TX |
| ZIP/Postal code |
75063 |
| Country |
USA |
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| Platform ID |
GPL16791 |
| Series (2) |
| GSE200992 |
The PNUTS-protein phosphatase 1 complex acts as an intrinsic barrier to KSHV replication [ChIP-seq] |
| GSE201046 |
The PNUTS-protein phosphatase 1 complex acts as an intrinsic barrier to KSHV replication |
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| Relations |
| BioSample |
SAMN27622633 |
| SRA |
SRX14894051 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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