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| Status |
Public on Jun 01, 2024 |
| Title |
CHiC_MD_E115-WT |
| Sample type |
SRA |
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| Source name |
Embryonic Mandible
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| Organism |
Mus musculus |
| Characteristics |
tissue: Embryonic Mandible
strain: FVB
embryonic day: E11.5
genotype: WT
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| Growth protocol |
Embryonic forelimbs, mandibular processes, and hearts of wildtype embryos at E11.5 (48-52 somite-stage) were microdissected in cold PBS and subjected to tissue dissociation.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Tissues were homogenized using a Dounce tissue grinder and fixed with 2% formaldehyde (37%) for 10 min. The reaction was quenched using 1.25M Glycine and pellets were snap-frozen for storage at -80°C. For nuclei isolation pellets were resuspended in custom-made lysis buffer (10mM Tris pH7.5, 10mM NaCl, 5mM MgCl2, 0.1mM EGTA, Protease Inhibitor). After on wash with PBS samples were digested with DpnII, re-ligated, de-crosslinked and purified.
The 3C-library was sheared and after adaptor ligation and amplification of sheared DNA fragments, libraries were hybridized to custom-designed SureSelect beads (SureSelectXT Custom 0.5-2.9Mb library) and indexed following Agilent’s instructions. Multiplexed libraries were sequenced using a Illumina HiSeq 4000 (50bp paired-end).
3C-library, hybridization to custom-designed SureSelect beads (Agilent)
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 4000 |
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| Data processing |
Sequencing was performed to have comparable depth among tissue samples and reads were mapped to the reference genome GRCm38/mm10, filtered and de-duplicated using HiCUP pipeline v0.8.1. The pipeline was set up with Bowtie2 v2.4.5 for short read mapping. Juicer command line tools v1.9.9 was used for the generation of binned contact maps from valid and unique read pairs with MAPQ ≥ 30. Normalization and diagonal filtering was performed using the Cooler v0.8.11 matrix balancing tool with options “--mad-max 5 --min-nnz 10 --min-count 0 --ignore-diags 2 --tol 1e-05 --max-iters 200 --cis-only”. For binning and normalization, only the genomic region chr3:65196079-68696078 (mm10) that represents the targeted interval was considered and read pairs mapping to this interval were retained and shifted by the offset of 65,196,078 bp using custom chrom.sizes files. Balanced maps were exported at 5kb resolution with corrected coordinates (transformed back to original values). Subtraction maps were directly generated from Cooler balanced HiCmaps using hicCompareMatrices tool (HiCExplorer v3.7.2) with option “--operation diff”. Hi-C maps were visualized from .cool files using pyGenomeTracks.
Normalized inter-domain insulation scores(Crane et al. 2015)and TAD boundaries were determined using FAN-C(Kruse et al., 2020)(v.0.9.26-b2-beta) with ‘fanc insulation -w 15000 25000 35000 50000 75000 100000 130000’ and‘fanc boundaries --min-score 0.21’, respectively, from input .cool matrices. Tissue-differential insulation scores and boundary values were calculated using ‘fanc compare -c difference’and‘fanc boundaries --min-score 0.21’.
For virtual Capture-C, to better visualize individual interactions of target regions, a 5kb viewpoint centered on the coordinates of each genomic element of interest was used. Capture-C-like profiles were generated as described (Paliou et al., 2019) from filtered unique read pairs (hicup.bam files) which also served as input for CHi-C maps. Read-pairs were mapped based on MAPQ ≥ 30 and relative position of the two reads inside and outside the viewpoint, respectively. After quantitation of reads outside of the viewpoint (per restriction fragment), read counts were distributed into 3kb bins with proportional distribution of read counts in case of overlap with more than one bin. Each binned profile was smoothened by averaging over a running window of 5 bins and scaled by 10^3 / sum of all its counts on chr3 (Paliou et al., 2019). Peak profiles were generated using custom Java code based on htsjdk v2.12.0 (https://samtools.github.io/htsjdk/). The viewpoint and neighboring +/-5kb regions were excluded from computation of the scaling factor. The following genomic interval was used as Virtual Capture-C viewpoint: chr3:66,978,288-66,983,288 (centering Shox2 TSS). BigwigCompare tool (deepTools v3.5.1) was further used to generate relative subtraction Capture-C-like profiles.
Assembly: GRCm38/mm10
Supplementary files format and content: hic output files containing binned contact maps from unique read pairs (MAPQ ≥ 30) were generated by Juicer command line tools v1.9.9 (Durand 2016).
Supplementary files format and content: cool files containing balanced contact matrices generated with the hicConvertFormat tool (HiCExplorer v3.7.2) from native .hic out-puts.
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| Submission date |
May 19, 2023 |
| Last update date |
Jun 01, 2024 |
| Contact name |
Marco Osterwalder |
| E-mail(s) |
olheartdev@gmail.com
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| Organization name |
Universiy of Bern
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| Department |
DBMR
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| Lab |
Osterwalder Lab
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| Street address |
Murtenstrasse 24
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| City |
Bern |
| ZIP/Postal code |
3008 |
| Country |
Switzerland |
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| Platform ID |
GPL21103 |
| Series (2) |
| GSE232881 |
A gene desert required for regulatory control of pleiotropic Shox2 expression and embryonic survival [C-HiC] |
| GSE232887 |
A gene desert required for regulatory control of pleiotropic Shox2 expression and embryonic survival |
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| Relations |
| BioSample |
SAMN35178264 |
| SRA |
SRX20437396 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7385430_Mandible_S2_L008_R1R2_001.cool.gz |
215.4 Kb |
(ftp)(http) |
COOL |
| GSM7385430_Mandible_S2_L008_R1R2_001.hicup.MAPQ30.hic |
669.4 Kb |
(ftp)(http) |
HIC |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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