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| Status |
Public on Jul 10, 2024 |
| Title |
PPGS_4_H3K27me3 |
| Sample type |
SRA |
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| Source name |
breast tumor
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| Organism |
Mus musculus |
| Characteristics |
tissue: breast tumor
cell line: CCN6KOT
cell type: Breast Cancer cell line
condition: Spindle Metaplastic
treatment: Control
fraction: IP
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| Treatment protocol |
treatment with puromycin
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| Growth protocol |
CCN6KOT primary Cells were growth in DMEM/F12 10%FBS and trasnduced with lentivirus of DNTCF4 construct and control followed by puromycin selection
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
samples were fixed with 1% formaldehyde, quenched with glycine, and frozen as a pellet.
The Diagenode ChIPmentation kit with dual indices was used, following the manufacturer’s protocol. Briefly, the cells were lysed and resuspended in 130ul of chromatin shearing solution. Shearing was done in a Covaris S2 instrument. A 15ul aliquot of the sheared chromatin was taken for assessment of the shearing size. The remaining sheared chromatin split for immunoprecipitation in the presence of H3K27me3 antibody (purchased Cell Signaling, provided by the investigator) and IgG as input. Four ug of each antibody was used per immunoprecipitation. Immunoprecipitation was done overnight at 4C with rotation. The immunoprecipitates were next washed before on-bead tagmentation at 37C for 10 minutes with agitation. The reaction was stopped by putting the samples on ice and additional washes, before processing for stripping, end filling and reverse crosslinking. qPCR was used to determine the optimal cycle number for enrichment PCR, according to the manufacturer’s protocol. The final amplified libraries were cleaned using AMPure XP beads at a 1.8:1 bead:DNA ratio, quantitated with Qubit HS dsDNA, and fragment size assessed on a TapeStation HS D1000 kit. The libraries were qPCR quantitated for pooling using the KAPA Illumina qPCR quantitation kit
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
CL Convert Conversion Software v4.0 was used to generate de-multiplexed Fastq files
TrimGalore(TrimGalore)(v0.4.5) and cutadapt(Martin)(v1.15) were used with the following parameters: –nextera -e 0.1 –stringency 6 –length 20 –nextseq 20
trimmed reads were aligned to mm10 with Bowtie2(v2.3.4.1) using default parameters with the exception of the following flags: -X 2000
Duplicate reads are marked with Picard(v2.20.2)
Alignments are filtered with samtools(v1.2) using the flags: -F4 -F8 -f3 -q10 -F1024
Alignments completely overlapping blacklisted regions(ENCODE Blacklist Regions)are removed with bedtools(v2.28.0)
Sample-wise peaks are called with macs2 (v2.1.2) with flags: –format BAMPE –gsize mm
Assembly: mm10
Supplementary files format and content: .bed peak files
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| Submission date |
Dec 19, 2023 |
| Last update date |
Jul 10, 2024 |
| Contact name |
Bioinformatics Core |
| E-mail(s) |
bioinformatics@umich.edu
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| Organization name |
University of Michigan
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| Street address |
BRCF Bioinformatics Core, 2800 Plymouth Rd
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| City |
Ann Arbor |
| State/province |
Mi |
| ZIP/Postal code |
48109-2800 |
| Country |
USA |
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| Platform ID |
GPL24247 |
| Series (2) |
| GSE250574 |
CCN6/WISP3 suppresses metaplastic breast carcinoma by antagonizing a WNT/b-catenin/EZH2 cascade [ChIP-seq] |
| GSE250575 |
CCN6/WISP3 suppresses metaplastic breast carcinoma by antagonizing a WNT/b-catenin/EZH2 cascade |
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| Relations |
| BioSample |
SAMN38932110 |
| SRA |
SRX22969934 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7981894_PPGS_4_H3K27me3_peaks.bed.gz |
518.2 Kb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
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