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| Status |
Public on May 15, 2016 |
| Title |
27Ac D4 |
| Sample type |
SRA |
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| Source name |
H3K27Ac at day 4 of M9-induced reprogramming
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| Organism |
Mus musculus |
| Characteristics |
cell type: tdMEFs at day 4 of reprogramming
chip antibody: H3K27ac (Millipore, 17-683, lot 2510209)
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| Treatment protocol |
M9-induced neural reprogramming was performed in M9 medium (50% Neural basal, 50% DMEM/F12/Glutamax, 1xN2, 1xB27 without vitamin A, 0.075% BSA, 0.1 mM nonessential amino acids, CHIR99021 3 mM, LDN193189 100 nM, A83-01 0.5 mM, Hh-Ag1.5 0.5 mM, retinoic acid 1 mM, SMER28 10 mM, RG108 10 mM, Parnate 2 mM, bFGF 10 ng/ml) at 5% O2 and 5% CO2 incubator at 37°C.
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| Growth protocol |
tdMEFs were cultured in MEF medium (Dulbecco’s modified Eagle’s medium supplemented with 10 % fetal bovine serum, 0.1 mM nonessential amino acids, and 2 mM Glutamax) in 5 % CO2 and 20 % O2 at 37 °C.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells at indicated time points were cross-linked by 1% formaldehyde, quenched with 0.125 M glycine. The chromatin DNA was extracted and sheared into 200-400 bp, and immuoprecipitated using a EZ-ChIP kit (Millipore, 17-371), and indicated antibodies.
Barcoded ChIP-Seq libraries were made from 1 to 10 ng of chromatin immunoprecipitated DNA (ChIP-DNA) using the NuGen Ovation Ultralow V2 kit. 13 cycles were used for PCR amplification of the libraries. Amplified libraries were checked for adapter artifacts by Agilent Bioanalyzer using DNA high sensitivity reagents and chips (Agilent). All libraries were quantified by qPCR using an Illumina Library Quantification kit (KAPA Biosystems). Libraries were pooled in equimolar amounts and run on the HiSeq 2500 sequencer with a single read 50 cycle run (Illumina).
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
base-calling was done using CASAVA 1.8.4.
For the histone marker ChIP-seq data: the alignment was done using bowtie (version 1.1.0) with human genome build mm10, reporting only reads with unique genome alignments. The result was then filtered by removed potential PCR duplications with with samtools (version 0.1.19)
Read Intensity of Histone markers were calculated with bedtools multicov, then normalized by the total number of reads for each samples, result were mered into one single file
Genome_build: mm10
Supplementary_files_format_and_content: histone marker excel file contains the normalized intensity around the TSS of genes (-2kb to +3kb)
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| Submission date |
Mar 07, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Sheng Ding |
| Organization name |
THE J DAVID GLADSTONE INSTITUTES
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| Department |
Gladstone Institute of Cardiovascular Disease
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| Street address |
1650 Owens Street
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| City |
SAN FRANCISCO |
| State/province |
California |
| ZIP/Postal code |
94158 |
| Country |
USA |
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| Platform ID |
GPL17021 |
| Series (2) |
| GSE78977 |
Genome-wide chromatin landscape of chemical-induced neural stem cell reprogramming from defined fibroblasts |
| GSE78978 |
CHIP-seq analysis for chemical-induced neural stem cells from defined fibroblasts |
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| Relations |
| BioSample |
SAMN04537346 |
| SRA |
SRX1618377 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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