 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Jan 22, 2024 |
| Title |
HiC_Flx1: Satb2 flx/flx DIV14 primary cortical cultures, 1h bic, rep1 |
| Sample type |
SRA |
| |
|
| Source name |
Cortex
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Cortex
cell type: DIV14 primary cortical cultures
genotype: Satb2 flx/flx
treatment: 1h bicucculine
|
| Treatment protocol |
Cottical neurons were treated with 20 µM NBQX for 16 h on DIV13. After silencing, neurons were stimulated with 50 µM biccuculine for 1h on DIV14 and then collected.
|
| Growth protocol |
Corical tissue was dissected at P0-P1, dissociated with Papain and titurated. Cells were plated in serum-containing medium (MEM, 1% horse serum, 0.1% Pen/Strep, 0.1 % sodium piruvate). After 2h, MEM was excahnged with serum-free feeding medium (Neurobasal-A Medium, containing B-27, 0.1% Pen/Strep and 2 mM Glutamine. Cells were treated with 5µM cytosine arabinoside on DIV3 to inhibit glial cell proliferation. At DIV4, one third of the medium was replaced by fresh serum-free feeding medium. Neurons were cultured for 14 days in 5% CO2 at 37°C.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
For in situ Hi-C, Arima-HiC kit was used, according to the manufacturer’s protocol. Briefly, 2.5x106 primary cortical neurons were fixed directly on the dish by addition of formaldehyde to a final concentration of 1 % for 10 min on RT. After washing twice with 1X PBS cells were detached using a cell lifter and collected in 1X PBS containing protease inhibitors. Cells were pelleted at 500 g for 10 min at 4 °C and pellets were directly snap-frozen in liquid nitrogen. After nuclei isolation and chromatin digestion, biotin was added and filled ends were ligated. After reverse crosslinking and purification, DNA was shared using a M220 Covaris sonicator at duty factor 12 %, PIP 75, CPB 200 for 60 s to achieve input fragment size of 400 bp. Additionally, DNA was size-selected using AMPure XP beads, according to Arima HiC kit manual.
Hi-C libraries were prepared using Swift Biosciences Accel-NGS 2S Plus DNA Library Kit according to Arima-HiC Kit User Guide (November 2018)
|
| |
|
| Library strategy |
Hi-C |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Paired-end sequencing reads were mapped to the mouse reference genome assembly (mm10), artefacts were filtered, and libraries were ICED normalized using HiC-Pro (v.2.11.4) at diffrent resolutions.
Differential compartment analysis was carried out by using dcHiC. Input file was created following dcHiC guidelines using HiC-Pro matrices at 100 kb.
TADs were computed at 25 kb resolution using SpectralTAD. Differential TAD boundaries between cKO and Flx Hi-C contact matrices were identified by TADCompare.
Chromatin loops were called by Mustache (v1.2.0) at 5 kb resolution using pooled replicate Hi-C matrices. The following parameters for differential loop detection were used: P value (-p) of 0.05 and sparsity (-st) of 0.8 thresholds together with KR normalization.
Frequently interacting regions (FIREs) were called using FIREcaller. Briefly, ‘sparseToDense.py’ HiC-Pro utility was used to create quadratic matrices at 10 kb resolution.
Assembly: mm10
Supplementary files format and content: bedgraph / genomic coordinates and EV1 values of called compartments in Flx and cKO samples
Supplementary files format and content: txt / differential TAD boundaries between cKO and Flx samples
Supplementary files format and content: bedpe / genomic coordinates of chromatin loops in Flx and cKO samples called 5kb resolution
Supplementary files format and content: txt / FIRE analysis in Flx and cKO samples
|
| |
|
| Submission date |
Jan 11, 2023 |
| Last update date |
Jan 22, 2024 |
| Contact name |
Nico Wahl |
| E-mail(s) |
nico.wahl@i-med.ac.at
|
| Phone |
0043512900371255
|
| Organization name |
Medical University Innsbruck
|
| Department |
Institute for Neuroscience
|
| Street address |
Innrain 66
|
| City |
Innsbruck |
| ZIP/Postal code |
6020 |
| Country |
Austria |
| |
|
| Platform ID |
GPL24247 |
| Series (2) |
| GSE222606 |
SATB2 organizes the 3D genome architecture of cognition in cortical neurons [Hi-C] |
| GSE222609 |
SATB2 organizes the 3D genome architecture of cognition in cortical neurons |
|
| Relations |
| BioSample |
SAMN32676504 |
| SRA |
SRX19002741 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
|
|
|
|
 |
