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| Status |
Public on Sep 27, 2019 |
| Title |
rB_wt input |
| Sample type |
SRA |
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| Source name |
resting B cell
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| Organism |
Mus musculus |
| Characteristics |
genotype: WT
tissue: spleen
chip antibody: input
activation time: 0h
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIP-Seq: Cultured cells were fixed with 1% formaldehyde (Sigma) for 10’ at 37°C. Fixation was quenched by addition of glycine (Sigma) at a final concentration of 125 mM. Twenty million fixed cells were washed with PBS and resuspended in 1 ml of RIPA buffer (10 mM Tris [pH 7.6], 1 mM EDTA, 0.1% SDS, 0.1% sodium deoxycholate, 1% Triton X-100, 1× Complete Mini EDTA free proteinase inhibitor (Roche)) or stored at −80°C until further processing. Sonication was performed using Covaris S2 sonicator at duty cycle 20%, intensity 5, cycle/burst 200 for 30 min or Branson sonifier at amplitude 35%, 12 cycles of 20” sonication and 30” of pause. For native chip, chromatin was digested with Mnase (Sigma) in digestion buffer (50 mM Tris-HCl, pH7.6, 1 mM CaCl2, 0.2% Triton X-100, butyrate 5 mM) for 5’ at 37°C and dialyzed against RIPA buffer for 2hrs at 4°C. Five micrograms of antibody were incubated with 40 μl of Dynabeads Protein A (or G) for 40 min at room temperature. Antibody-bound beads were added to 500 μl of sonicated or Mnase-digested chromatin, incubated at 4°C overnight, and washed twice with RIPA buffer, twice with RIPA buffer containing 0.3M NaCl, twice with LiCl buffer (0.25 M LiCl, 0.5% Igepal-630, 0.5% sodium deoxycholate), once with TE (pH 8.0) plus 0.2% Triton X-100, and once with TE (pH 8.0). Crosslinking was reversed by incubating the beads at 65°C for 4 hr in the presence of 0.3% SDS and 1 mg/ml Proteinase K. ChIP DNA was purified by phenol-chloroform extraction followed by ethanol precipitation.
ChIP-Seq : Library was prepared in Ovation SP Ultralow library system (Nugen). 50 cycles of sequencing data were acquired on the HiSeq 2000 (Illumina).
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
input
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| Data processing |
base calling: illumina CASAVA 1.8.2
ChIP-Seq_density: alignment: bowtie-1.1.1/bowtie -S -m 1 -p 20 -a --best --strata -n 2 -l 50
ChIP-Seq_density: filtering (uniquely aligned reads): samtools-0.1.19-1.2/samtools view -S -b -F4
ChIP-Seq_density: normalized tag density: custom script, maximum 2 reads with the same start position allowed
Genome_build: mm9
Supplementary_files_format_and_content: ChIP-Seq: .wig files tag density in 100 nt windows divided by windowsize and library size in millions to obtain normalized read density (rpkm)
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| Submission date |
Sep 26, 2019 |
| Last update date |
Sep 27, 2019 |
| Contact name |
Seolkyoung Jung |
| Organization name |
NIH
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| Department |
NIAMS
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| Lab |
biodata mining and discovery section
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| Street address |
10 Center Dr
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| City |
bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
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| Platform ID |
GPL13112 |
| Series (1) |
| GSE82144 |
Myc regulates chromatin decompaction and nuclear architecture during B cell activation |
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| Relations |
| BioSample |
SAMN12857648 |
| SRA |
SRX6912441 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4098725_rB_wt_input.bw |
42.3 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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