|
| Status |
Public on Jan 01, 2015 |
| Title |
naiveT-input_H3K4me3 |
| Sample type |
SRA |
| |
|
| Source name |
spleen
|
| Organism |
Mus musculus |
| Characteristics |
cell type: CD4+
chip antibody: none
genetic background: C57bl/6
|
| Treatment protocol |
For purification of activated cells, naïve CD4+ T cells were prepared as described. Plates were coated with goat-anti-hamster antibodies followed by anti-CD3e (2C11) and the naïve T cells plated in complete RPMI including anti-CD28. 72 hours later cells were harvested, fixed with formaldehyde and frozen.
|
| Growth protocol |
CD4+ T cells were isolated from spleens from C57bl/6 mice by magnetic bead selection.Cells were treated with biotinylated antibodies allowing a negative selection on the Automacs separation system (Miltenyi Biotech). Cells were fixed in formahdehyde and frozen. Purity of CD4 T cells was greater than 90%.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated cells and protein-DNA complexes were isolated with antibody. Libraries were prepared according to Illumina's instructions. Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (minus exo) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation, ChIP DNA was PCR-amplified with Illumina primers with the respective indexes for 18 cycles and library fragments (ChIP) (insert plus adaptor and PCR primer sequences) were size- selected from a 8% polyacrylamide gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on a HiSeq 2000 (Illumina) following the manufacturer's protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
For ChIP-Seq (single end reads), reads were aligned to the mm9 genome using Bowtie (v0.12.7), keeping only reads that mapped to a single, unique location. Aligned read files were analyzed with HOMER (http://biowhat.ucsd.edu/homer/).
Genome_build: mm9
Supplementary_files_format_and_content: Processed files include bedgraph files (Chipseq fragment density files which were analyzed using the UCSC Genome Browser) and a bed file (peak position file used for motif analysis using HOMER)
|
| |
|
| Submission date |
Apr 30, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Carol Katayama |
| E-mail(s) |
ckatayama@ucsd.edu
|
| Phone |
858-534-5961
|
| Fax |
858-534-0980
|
| Organization name |
UC San Diego
|
| Department |
BIO/CMM
|
| Lab |
Hedrick
|
| Street address |
9500 Gilman Dr.
|
| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92024 |
| Country |
USA |
| |
|
| Platform ID |
GPL13112 |
| Series (1) |
| GSE46525 |
ICOS coreceptor signaling inactivates the transcription factor FOXO1 to promote Tfh cell differentiation |
|
| Relations |
| BioSample |
SAMN02087668 |
| SRA |
SRX272972 |
