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| Status |
Public on Dec 21, 2022 |
| Title |
Pt2a Hi-C rep 1 |
| Sample type |
SRA |
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| Source name |
Pt2a
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| Organism |
Pan troglodytes |
| Characteristics |
cell line: Pt2a
cell type: iPS-derived neural progenitor cells
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| Treatment protocol |
Hi-C was performed using the Arima Hi-C kit (Arima Genomics) according to the manufacturer’s instructions. 10 million cells were used. The sequencing library was prepared using Accel-NGS 2S Plus DNA Library Kit (Swift Biosciences) according to the manufacturer's protocol. Two independent biological replicates were prepared for each cell line. In total eight libraries were pooled and sequenced with paired-end 150-bp reads using two lanes of a NovaSeq6000 S2 (Illumina) at the Chan Zuckerberg Biohub.
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| Growth protocol |
Two human (WTC11 and HS1) and two chimpanzee (C3649 and Pt2a) induced pluripotent cell lines (iPSCs) were cultured in Matrigel-coated plates with mTeSR media (WTC11 and C3649 were cultured in StemFlex) in an undifferentiated state. Cells were propagated at a 1:3 ratio by treatment with 200 U/mL collagenase IV (or PBS-EDTA) and mechanical dissection. WTC11 and C3649 iPSCs were differentiated to neural progenitor cells (NPCs) as previously described (64). Briefly, 2-2.5×10⁵ cells per cm² were seeded on Matrigel-coated wells in StemFlex containing 2 μM Thiazovivin. The following day (Day 0), medium was replaced with E6 containing 500 nM LDN193189 (Selleckchem), 10 μM SB431542 (Selleckchem), and 5 μM XAV-939 (Selleckchem). Starting on Day 3, medium was replaced with E6 containing 500 nM LDN193189 and 10 μM SB431542 every 48 hrs. Starting on Day 12, medium was replaced with Neurobasal containing 2 mM GlutaMAX, 60 μg per ml L-Ascorbic acid 2-phosphate, N2, and B27 without Vitamin A every 48 hours. Around Day 16, cells were washed with PBS, dissociated with Accutase, pelleted and resuspended in Neurobasal containing 2 mM GlutaMAX, 60 μg per ml L-Ascorbic acid 2-phosphate, N2, and B27 without Vitamin A, 10 ng per ml fibroblast growth factor 2, and 10 ng per ml epidermal growth factor, and seeded on poly-L-ornithine-, fibronectin-, and laminin-coated wells. Cells were collected for HiC at passage 5-7. To differentiate HS1 and Pt2a iPSCs into NPCs, cells were split with EDTA at 1:5 ratios in culture dishes coated with matrigel and culture in N2B27 medium (comprised of DMEM/F12 medium (Invitrogen) supplemented with 1% MEM-nonessential amino acids (Invitrogen), 1 mM L-glutamine, 1% penicillin-streptomycin, 50 ng/mL bFGF (FGF-2) (Millipore), 1x N2 supplement, and 1 x B27 supplement without Vitamin A (Invitrogen)) supplemented with 100 ng/ml mouse recombinant Noggin (R&D systems). Cells at passages 1-3 were split by collagenase into small clumps, and continuously cultured in N2B27 medium with Noggin. After passage 3, cells were plated at the density of 5×10⁵ cells/cm² after disassociation by TrypLE express (Invitrogen) into single-cell suspension, and cultured in N2B27 medium supplemented with 20 ng/mL bFGF and EGF. Cells were maintained and collected at passage 18-20. Our use of two differentiation protocols reflects rapid progress in stem cell research during the course of this study.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
The sequencing library was prepared using Accel-NGS 2S Plus DNA Library Kit (Swift Biosciences) according to the manufacturer's protocol. Two independent biological replicates were prepared for each cell line. In total eight libraries were pooled and sequenced with paired-end 150-bp reads using two lanes of a NovaSeq6000 S2 (Illumina) at the Chan Zuckerberg Biohub.
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| Library strategy |
Hi-C |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
Adapters were trimmed from raw FASTQ files using TrimGalore [v0.6.5] with options --illumina --paired.
The data were then processed from adapter-trimmed FASTQ files to Hi-C contacts as cooler files using Distiller [v0.3.3] (65). This processing includes read mapping with BWA-MEM (66), filtering (MAPQ >= 30), contact pair processing with pairtools (67) and normalization via matrix balancing (68). Samples were processed both per replicate, per individual and per species. For easier comparison of samples in some analyses, we mapped the data from each species to the reference genome of the other species (human to pantro6 and chimp to hg38).
Genome_build: hg38, panTro6
Supplementary_files_format_and_content: Cooler (Hi-C contact matrices)
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| Submission date |
Aug 31, 2021 |
| Last update date |
Dec 21, 2022 |
| Contact name |
Katherine S Pollard |
| E-mail(s) |
katherine.pollard@gladstone.ucsf.edu
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| Organization name |
Gladstone Institutes
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| Department |
Institute of Data Science and Biotechnology
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| Street address |
1650 Owens St
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| City |
San Francisco |
| State/province |
CA |
| ZIP/Postal code |
94158 |
| Country |
USA |
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| Platform ID |
GPL30573 |
| Series (1) |
| GSE183137 |
Three-dimensional genome re-wiring in loci with Human Accelerated Regions |
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| Relations |
| BioSample |
SAMN21161154 |
| SRA |
SRX11977907 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5551872_chimp_Pt2a_rep1.panTro6.mapq_30.1000.mcool |
1.5 Gb |
(ftp)(http) |
MCOOL |
| GSM5551872_chimp_Pt2a_rep1_hg38.hg38.mapq_30.1000.mcool |
1.0 Gb |
(ftp)(http) |
MCOOL |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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