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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Jul 26, 2024 |
| Title |
NPC, 32°C, replicate 1 |
| Sample type |
SRA |
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| Source name |
Brain
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| Organism |
Mus musculus |
| Characteristics |
tissue: Brain
cell line: cortical mNPC
cell type: nouse neural progenitor cells
strain: C57Bl/6NTac
treatment: 32degreeC 6h
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| Treatment protocol |
Following 36 h of incubation at 37°C, one plate with 4 lines was incubated at 32°C for 6 h.
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| Growth protocol |
Mouse NPCs were isolated from the cortex of E17.5 C57BL6/NTac embryos. The protocol was adapted from Bernas et al, 2017105. Embryo cortices were manually dissected from brains and the tissue was dissociated in 1X TrypLE™ Select Enzyme (Thermo Fisher Scientific, A1217701) for 10 min, with manual dissociation using a 1000 µL pipette. The cell suspension was washed in Neurobasal medium (Thermo Fisher Scientific, 21103049) and filtered through a 70 µm cell strainer (Miltenyi Biotech, 130-110-916). Cells were washed twice in 5 mL Neurobasal medium (Gibco, 21103049) by centrifugation at 200 xg for 10 min. Cells were resuspended in 1 mL Neurobasal growth medium containing 1X B27 supplement (Thermo Fisher Scientific, 17504044), 1X Penicillin/Streptomycin (Thermo Fisher Scientific, 15140122), 1X Glutamax (Thermo Fisher Scientific, 35050038), 20ng/mL FGF-2 (Peprotech, 100-18B), 20 ng/mL EFG (Peprotech, AF-100-15), and 2 µg/mL Heparin (MP Biomedicals, 210193125). Cells were seeded onto 12-well plates, previously coated for 2 h at 37°C in 1:100 dilution of Matrigel (Corning, 354234). After passaging, NPCs were cultured in media described above on plates coated overnight in poly-D-lysine (Sigma, P7280), and then coated for 2 h in laminin (Sigma, L2020). Cells were either sub-cultured or media was changed every 2 days. Primary mouse NPC lines from 4 separate embryos were seeded 0.3 x 106 cells per well, in two batches, in a poly-D-lysine/laminin coated 6-well plate after 9 days in vitro.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Cells from both 37°C and 32°C conditions were harvested in tri-reagent and RNA was isolated using the Direct-zol RNA Microprep kit (Zymo Research, R2063).
RNAseq library construction and Illumina next generation sequencing was performed by Novogene on a PE150 flow cell, generating 30 million paired-end reads per sample.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
mouse_NPC_temp_DESeq.csv
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| Data processing |
Fastq files were pseudo-aligned to the GRCm39 mouse transcriptome, downloaded from NCBI, using Kallisto (v.0.48.0) with paired-end mode on and 100 bootstraps.
Kallisto output files were imported into R using the tximport package. Transcripts were assigned to genes using the TxDb.Mmusculus.UCSC.mm10.ensGene package (v.3.4.0) in R . Differential expression analysis was performed using the DESeq2 package (v.3.16) in R after discarding transcripts with less than 10 counts. Differentially expressed genes were defined as those with an adjusted p-value less than 0.1.
Assembly: mm39, downloaded from NCBI
Supplementary files format and content: HDF5 binary file containing run info, abundance esimates, bootstrap estimates, and transcript length information length.
Supplementary files format and content: csv file containing result from DESeq such as; count and p-value and logfoldchange
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| Submission date |
Jun 12, 2023 |
| Last update date |
Jul 26, 2024 |
| Contact name |
Hans Tomas Bjornsson |
| E-mail(s) |
htb@hi.is
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| Organization name |
University of Iceland
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| Department |
Faculty of Medicine
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| Lab |
Louma G. Laboratory of Epigenetic Research
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| Street address |
Sturlugata 8
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| City |
Reykjavik |
| State/province |
Hofudborgarsvaedid |
| ZIP/Postal code |
102 |
| Country |
Iceland |
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| Platform ID |
GPL24247 |
| Series (2) |
| GSE234688 |
SMYD5 is a regulator of the mild hypothermia response [RNA-Seq] |
| GSE234699 |
SMYD5 is a regulator of the mild hypothermia response |
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| Relations |
| BioSample |
SAMN35715776 |
| SRA |
SRX20661614 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7473349_NPC2194_3_32_20230102_Kallisto_abundance.h5 |
50.1 Mb |
(ftp)(http) |
H5 |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
| Processed data provided as supplementary file |
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