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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Jun 07, 2018 |
| Title |
Hi-C D4 EB |
| Sample type |
SRA |
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| Source name |
mouse embryonic stem cells
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| Organism |
Mus musculus |
| Characteristics |
cell type: mouse embryonic stem cells
genotype: WT
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| Growth protocol |
ES cell–derived neural progenitor cell clones were cultured on poly-L-ornithine (0.001%) and laminin (1μg/ml) coated plates with N2 media (DMEM/F-12, GlutaMAX™ supplement, 1X N2 supplement, 1X B27 supplement, 1X Pen/Strep) supplemented with FGF2 (20ng/ml), EGF (20ng/ml), laminin (1μg/ml), and heparin (10μg/ml). Undifferentiated ES cells were grown on 0.2% gelatin-coated plates with N2B27 media (mixture of 500ml DMEM/F12 media, 500ml Neurobasal media, 5ml N2 supplement, 10ml B27 supplement, 10ml Pen/Strep, 5ml GlutaMAX, and 2ml 55mM β-mercaptoethanol) supplemented with 1μM PD0325901, 3μM CHIR-99021, and 1000U/ml LIF. Embryoid bodies (EBs) were grown with differentiation media (DMEM, high glucose, GlutaMAX supplement, pyruvate, 15% Hyclone FBS, 25mM HEPES pH 7.2-7.5, 1X MEM non-essential amino acids, 1X Pen/Strep, 0.1mM β-mercaptoethanol) in suspension (D4 EBs) or on gelatin-coated plates (D7 EBs).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Total RNA was extracted using standard Trizol protocol. ChIP/CHART-enriched and input DNA were extracted with phenol:chloroform.
RNA-seq libraries were generated by dUTP-mediated directional RNA-seq based on NEBNext Ultra directional RNA library preparation protocol. ChIP-seq and Hi-C libraries were generated by NEBNext ChIP-Seq Library Prep Master Mix Set for Illumina. CHART-seq libraries were generated by NEBNext Ultra DNA Library Prep Kit for Illumina.
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| Library strategy |
Hi-C |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
In situ Hi-C: Each end of the read-pairs was aligned separately as single-end reads to 129S1/SvJm (mus) and CAST/Eih (cas) genome using NovoAlign_v3-1.00.02 and allele-specificity was assigned. The single-end alignments were joined to generate a Hi-C summary file.
Genome_build: mm9
Hi-C: Allele-resolved Hi-C data were presented as Hi-C summary file (file names ending with '.summary.txt'), where the first 7 columns represent read names, chromosome for read 1, positions for read 1 (5' end of read), strand of read 1 (+ or -), chromosome for read 2, positions for read 2 (5' end of read), and strand of read 2 (+ or -), respectively. WT1, WT clone 1. KO1, Smchd1-/- clone 1. rep1, replicate 1. rep2, replicate 2. cas, cas-specific reads. mus, mus-specific reads. comp, all reads. Two biological replicates were pooled and principle component analysis was performed on pooled Hi-C data at 200-kb resolution using HiTC. PC1 values were presented as bedGraph files (file names ending with '.PC1.bedGraph'). WT1, WT clone 1. KO1, Smchd1-/- clone 1. D0, undifferentiated ES cells. D4, day4 differentiated embryoid bodies. D7, day7 differentiated embryoid bodies. Xa, PC1 values derived from Xcas reads (WT1, KO1, D4, and D7) or all reads mapped to the X (D0). Xi, Xmus reads. To match sequencing depths, 50% reads from WT1 rep1 and KO1 rep1 Hi-C summary files were randomly sampled to generate down-sampled Hi-summary files and PC1 profiles (file names starting with 'Downsampled'). We also reanalyzed published undifferentiated female ES cells Hi-C data (GSE72697: GSM1868575). Reads were aligned to the mm9 genome using bowtie2 and the PC1 values of the X chromosome at 200-kb resolution were similarly derived using total X chromosomal reads (Giorgetti.D0.HiC.Xa.comp.PC1.bedGraph). Insulation scores were computed using iteratively corrected Hi-C contact matrices at 40-kb resolution as described (Giorgetti et al., 2016) and summarized in X_chromosome.insulation_score.txt.
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| Submission date |
Mar 09, 2018 |
| Last update date |
Apr 15, 2019 |
| Contact name |
Chen-Yu Wang |
| E-mail(s) |
cywang@molbio.mgh.harvard.edu
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| Organization name |
Massachusetts General Hospital
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| Department |
Molecular Biology
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| Lab |
Jeannie T. Lee
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| Street address |
185 Cambridge Street
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| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02114 |
| Country |
USA |
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| Platform ID |
GPL17021 |
| Series (1) |
| GSE99991 |
Investigating the role of SMCHD1 in de novo X chromosome inactivation |
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| Relations |
| Reanalyzed by |
GSM3722628 |
| BioSample |
SAMN08683055 |
| SRA |
SRX3779313 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3036557_D4.HiC.cas.summary.txt.gz |
145.1 Mb |
(ftp)(http) |
TXT |
| GSM3036557_D4.HiC.comp.summary.txt.gz |
1.8 Gb |
(ftp)(http) |
TXT |
| GSM3036557_D4.HiC.mus.summary.txt.gz |
671.1 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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