|
| Status |
Public on Sep 30, 2021 |
| Title |
H3K7me3_ChIPSeq rep 3 |
| Sample type |
SRA |
| |
|
| Source name |
resting B cells isolated from spleen
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
genotype: WT
cell type: Resting B cells
chip antibody: H3K27me3 (NEB- 9733S)
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Spleens were isolated from 6- to 10-week-old C57B6 mice and homogenized through a sieve in B cell culture medium (RPMI-1640 (Lonza), 10% fetal calf serum (FCS) (Sigma), 0.1 U/ml penicillin (Lonza), 0.1 μg/ml streptomycin (Lonza), 2mM L-Glutamine (Lonza), 50μM beta-mercaptoethanol (Gibco)). The cell suspension was centrifuged on a Ficoll-Paque (GE Healthcare) cushion and the buffy coat layer was resuspended at a concentration of 1x 10e8 cells/ml in PBS + 2% FCS/ 1mM EDTA. Resting B cells were isolated using an EasySep Negative Selection- Mouse B Cell Isolation Kit (STEMCELL Technologies), which depletes for non-B cell markers and activated CD43+ B cells.
5ng of ChIP/Input DNA was used to prepare libraries using the NEBNext® Ultra™ II DNA LibraryPrep Kit for Illumina following the manufacturer recommendations. Library quality and quantity were assessed on a Bioanalyser and Qubit, respectively. Libraries were sequenced on an Illumina Hiseq 2500 (v4 chemistry), and single-end 50bp reads or paired-end 100bp reads per sample were generated.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
H3K27me3 PS3
Three biological replicates H3K27me3 PS1, H3K27me3 PS2 and H3K27me3 PS3 were merged for downstream analysis. More details please see the Data processing section.
H3K27me3_CPM.bw
|
| Data processing |
Illumina CASAVA 1.8.4 software used for basecalling.
Raw reads were aligned to UCSC mm9 mouse genome obtained from iGenome with BWA (0.7.5a) for single-end reads and bowtie2 (2.3.4) with arguments -p 8 --local --very-sensitive-local --no-unal --no-mixed --no-discordant --phred33 -I 10 -X 700 for paired-end reads.
Aligned reads were sorted and converted to bam by samtools (1.6).
Duplicated reads were removed by Picard MarkDuplicates tool (1.9).
Merged bam files from biological replicates were produced by samtools.
Duplicated reads were removed from merged bam files by Picard MarkDuplicates tool (1.9).
bigWig files were generated from merged duplicated reads removed bam files by bedtools genomeCoverageBed (normalized to Reads Per Million; RPM) and bedGraphToBigWig obtained from UCSC.
Genome_build: mm9 (MGSCv37)
Supplementary_files_format_and_content: bigWig files
|
| |
|
| Submission date |
Dec 04, 2020 |
| Last update date |
Sep 30, 2021 |
| Contact name |
LMS Bioinformatics Core |
| E-mail(s) |
bioinformatics@lms.mrc.ac.uk
|
| Organization name |
MRC London Institute of Medical Sciences
|
| Department |
Bioinformatics Core
|
| Street address |
Hammersmith Hospital Campus, Du Cane Road
|
| City |
London |
| ZIP/Postal code |
W12 0NN |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL17021 |
| Series (2) |
| GSE162703 |
Genome-wide maps of transcription factor binding and histone modifications in splenic resting B cells isolated from mice |
| GSE162704 |
RUNX1 controls the rate of cell cycle entry during naive B cell activation by regulating expression of cell cycle and immunomodulatory genes in response to BCR stimulation |
|
| Relations |
| BioSample |
SAMN17004682 |
| SRA |
SRX9633973 |
