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| Status |
Public on Nov 02, 2017 |
| Title |
hct116.input.del.mll4.set |
| Sample type |
SRA |
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| Source name |
HCT116
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| Organism |
Homo sapiens |
| Characteristics |
cell line: HCT116
genotype/variation: MLL4-SET-KO
shRNA: NA
chip antibody: ChIP-Input
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| Treatment protocol |
Cells were cultures in DMEM containing 10% FBS
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| Growth protocol |
Cells were cultures in DMEM containing 10% FBS
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
RNA was extracted using Trizol reagent (Life technologies). RNAse free DNaseI (Sigma) was used to eliminate DNA contamination and the treated RNA was purified with RNeasy mini kit (Qiagen).
Cells were fixed with 1% formaldehyde for 10 minutes. After quenching and cell lysis, chromatin were sonicated using a E220 focused-ultrasonicator (Covaris). 100 µg sheared chromatin, 5 µg antibody, and 50 µl protein A/G beads (Santa Cruz) were used for each immunoprecipitation. Immunoprecipitated DNA were purified after washing, eluting, and reverse-croslinking and submitted for library preparation.
ChIP-seq libraries were prepared using the KAPA HTP Library Preparation Kit complemented with NEXTflex DNA Barcodes from Bioo Scientific. 10 ng of DNA was used as starting material for input and ip samples. Libraries were amplified using 13 cycles on the thermocycler. Post amplification libraries were size selected at 250- 450bp in length using Agencourt AMPure XP beads from Beckman Coulter. Libraries were validated using the Agilent High Sensitivity DNA Kit. RNA-seq libraries were prepared using the Illumina TruSeq Stranded Total RNA Preparation Kit with Ribo-Depletion. Input RNA quality was validated using the Agilent RNA 6000 Nano Kit. 200ng-1 µg of total RNA was used as starting material. Libraries were validated using the Agilent DNA 1000 Kit
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
Basecalls were performed using bcl2fastq v2.17 for NextSeq output.
Both ChIP-seq and RNA-seq reads were trimmed from 3' end until the final base had a quality score > 30, using Trimmomatic v0.33, discarding reads left with < 20 bp
ChIP-seq reads were aligned to the UCSC mm9, hg19, and dm3 genomes using Bowtie version 0.12.9. Only uniquely mapping reads with at most two mismatches were retained.
RNA-seq reads were aligned to the UCSC hg19 and dm3 genomes using Tophat version 2.0.9. Only uniquely mapping reads with at most two mismatches were retained.
To create ChIP-seq coverage plots, the locations of the mapped ChIP-seq reads were extended to 150 bp to represent sequenced fragments, renormalized (to reads per million, rpm) and reformatted in the bigWig file format.
To create RNA-seq coverage plots, the mapped RNA-seq reads were split according to strand, renormalized (to reads per million, rpm) and reformatted in the bigWig file format.
Genome_build: dm3, mm9, hg19
Supplementary_files_format_and_content: bigWig files were generated using the genomicRanges R package. Score represents the normalized coverage of RNA or DNA fragments at a given genomic coordinate.
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| Submission date |
Jul 19, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Ali Shilatifard |
| E-mail(s) |
[email protected]
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| Organization name |
Northwestern University Feinberg School of Medicine
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| Department |
Department of Biochemistry and Molecular Genetics
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| Lab |
Shilatifard Lab
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| Street address |
320 E Superior St
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| City |
Chicago |
| State/province |
IL |
| ZIP/Postal code |
60611 |
| Country |
USA |
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| Platform ID |
GPL18573 |
| Series (1) |
| GSE101646 |
A cryptic Tudor domain links BRWD2/PHIP to COMPASS-mediated histone H3K4 methylation |
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| Relations |
| BioSample |
SAMN07370736 |
| SRA |
SRX3020584 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2711400_hct116.input.del.mll4.set.bw |
302.5 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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