|
| Status |
Public on Jan 01, 2016 |
| Title |
input DNA_ChIPseq |
| Sample type |
SRA |
| |
|
| Source name |
embryonic stem cells
|
| Organism |
Mus musculus |
| Characteristics |
strain: Stella:GFP
genotype: wild type
antibody: none
|
| Growth protocol |
For maintenance all mouse ESC lines were grown in so-called “2i+LIF” medium that is serum free N2B27-based medium supplemented with MEK inhibitor PD0325901 (0.8 μM) and GSK3β inhibitor CHIR99021 (3.3 μM) in tissue culture (TC) dishes pretreated with 7.5 μg/ml polyL-ornithine (Sigma) and 5 μg/ml laminine (BD) (Hayashi and Saitou, 2013; Hayashi et al., 2011). For ChIP-seq, RNA-seq and immunoprecipitation experiments Stella:GFP mESC lines and FH-Prdm14 derivatives were cultured for 5 days in feeder-free conditions in DMEM-high glucose medium (DMEM/high glucose; HyClone) containing 15% (v/v) FBS, 1 mM glutamine, penicillin/streptomycin, 0.1mM nonessential amino acids, 0.1 mM 2-mercaptoethanol and supplemented with LIF (serum+LIF conditions).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIPs were performed using approximately 1-2 x 10^7 mESCs fixed with 1% formaldehyde for 10 min at room temp. RNA was extracted using Trizol.
Sequencing libraries were prepared starting from ~30ng of ChIP DNA or oligo-dT purified mRNA using the NEBNext Multiplex Oligos for Illumina kit (Cat# E7335S).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Reads aligned to mm9 using default settings with Bowtie2.2.4
Wig files were generated with QuEST2.4
RNA-seq were aligned to mm9 and quantified against mouse ENSEMBL 78 (GRCm38) gene models using htseq-count. Variance-stabilizing transformation (VST) was applied across samples for processed data.
Genome_build: Alignments done using mm9 as reference
Supplementary_files_format_and_content: wig and tab delimited text
|
| |
|
| Submission date |
Oct 30, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Natalie Nady |
| Organization name |
Stanford
|
| Department |
Chemical and Systems Biology
|
| Lab |
Wysocka
|
| Street address |
265 Campus Dr.
|
| City |
Stanford |
| State/province |
CA |
| ZIP/Postal code |
94303 |
| Country |
USA |
| |
|
| Platform ID |
GPL19057 |
| Series (1) |
| GSE74547 |
ETO Family Protein Mtgr1 Mediates Prdm14 Functions in Stem Cell Maintenance |
|
| Relations |
| BioSample |
SAMN04227577 |
| SRA |
SRX1404244 |
