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| Status |
Public on Nov 12, 2024 |
| Title |
chimpanzee neurons, differentiated, C3649, 2-week, rep1 (ATAC-seq) |
| Sample type |
SRA |
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| Source name |
C3649
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| Organism |
Pan troglodytes |
| Characteristics |
cell line: C3649
cell type: excitatory neurons
genotype: WT
treatment: differentiation
time: 2 week
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| Treatment protocol |
Human and chimp excitatory neurons were generated by integration of the AAVS1 safe harbor locus of human iPSCs (i3N iPSCs). Based on the similarity (99%) of conserved sequence of AAVS1 between human and chimpanzee genome, the same iNgn2 cassette used in human iPSCs was knocked in chimpanzee AAVS1 safe harbor locus. The iPSCs were dissociated and seeded in Matrigel coated plate in N2 medium containing BDNF (PeproTech, #450-02), NT-3 (PeproTech, #450-03), mouse laminin (Lifetechnology, #23017-015), Doxycycline (Sigma, #D9891) and Rock Inhibitor (STEMCELL Technologies, 72302), and changed medium for the following two days without Rock Inhibitor. After three days in N2 medium, the pre-differentiated cells were dissociated and seeded in a Poly-L-Ornithine coated plate in Maturation medium containing those above factors except Rock Inhibitor. For the following differentiation procedures, half medium change was applied for the cells every 7 days.
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| Growth protocol |
A well-characterized human iPSC line with a wildtype genetic background (WTC11) and a 2nd human iPSC F12468 iPSC line with the Ngn2 transgene were used for human cell culture. For chimpanzee cell culture, the iPS cell lines (C3649 and C3624), a gift from Yoav Gilad’s Lab, were used. iPSC lines were maintained on Matrigel-coated (Corning, 354277) plates with fresh mTeSR medium (StemCell Technologies, 5850) every two days. Both human and chimpanzee iPSCs were passaged using Accutase (STEMCELL Technologies, 07922) and mTeSR medium containing 10uM Rock inhibitor Y-27632 (STEMCELL Technologies, 72302). Human embryonic kidney (HEK) 293T cells were cultured in Dulbecco’s modified Eagle’s medium (Gibco, 11995065) supplemented with 10% fetal bovine serum (CPS Serum, FBS-500). HEK 293T cells were passaged with trypsin-EDTA (Gibco, 25200072). All cells were grown at 37°C with 5% CO2. Culture medium was routinely assessed for mycoplasma contamination with the MycoAlert Mycoplasma Detection Kit (LONZA, LT07-218).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Generally, live cells were washed once with ice-cold 1xPBS containing protease inhibitor then resuspended in ice-cold nuclei extraction buffer (10 mM Tris-HCl pH 7.5, 10 mM NaCl, 3 mM MgCl2, 0.1% Igepal CA630, and 1× protease inhibitor) for 5 min. Next, counted 50,000 cells per reaction and exchanged into 50 μL 1× buffer TD, and added 2.5 μL TDE1 enzyme for 30 min at 37 °C with rotation for transposition. The fully transposed DNA was purified with Qiagen MinElute spin columns (Qiagen#28006), PCR amplified, and AMPure XP beads size-selected for fragments between 300 and 1,000 bp. Libraries were subjected to paired-end 150 bp sequencing on the Novaseq 6000.
ATAC-seq was performed by using the Nextera DNA Library Prep Kit (Illumina FC-121-1030).
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| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
raw_pantro5_2percent-idr.conservative_peak.narrowPeak.gz
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| Data processing |
All libaries were processed using the ENCODE ATAC-seq pipeline (v1.10.0) using default settings and idr cutoff at 2%
Briefly, reads were mapped to bowtie2, duplicates removed by picard and peaks were called by MACS2
Assembly: hg38, pantro5
Supplementary files format and content: bigwig, idr conserivative at 2%
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| Submission date |
Dec 27, 2022 |
| Last update date |
Nov 17, 2024 |
| Contact name |
Yin Shen |
| E-mail(s) |
Yin.Shen@ucsf.edu
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| Phone |
4155023403
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| Organization name |
UCSF
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| Department |
Department of Neurology
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| Lab |
Shen Lab
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| Street address |
513 Parnassus Ave
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| City |
SAN FRANCISCO |
| State/province |
CALIFORNIA |
| ZIP/Postal code |
94143-0410 |
| Country |
USA |
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| Platform ID |
GPL30573 |
| Series (2) |
| GSE221830 |
Comparative characterization of human accelerated regions in neurons [ATAC-seq] |
| GSE222113 |
Comparative characterization of human accelerated regions in neurons |
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| Relations |
| BioSample |
SAMN32421877 |
| SRA |
SRX18859775 |
| 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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