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Sample GSM8047578 Query DataSets for GSM8047578
Status Public on Jul 05, 2024
Title Hippocampus, post-onset 1, snRNAseq
Sample type SRA
 
Source name Hippocampus
Organism Mus musculus
Characteristics tissue: Hippocampus
age: Adult
Sex: F
genotype: Scn8aD/+
batch: 1
Extracted molecule nuclear RNA
Extraction protocol snRNA-seq sample collection was performed on fresh tissue over two batches: batch 1 (pre-onset 1 and control 1) and batch 2 (pre-onset 2&3 and control 2&3)At postnatal day 50 (n = 3 each), mice were euthanized and their brains were removed (Fig 1A-B). Bilateral hippocampi were immediately dissected and placed in a Dounce homogenizer with 1 mL lysis buffer: EZ Prep Lysis Buffer from Nuclei EZ Prep (Sigma #NUC101), 1mM DTT, 27U/mL Protector RNase inhibitor (Sigma #3335399001). Hippocampi were homogenized as follows: 25X with pestle A, 25X with pestle B, wait 2.5 minutes, 15X with pestle B, wait 2.5 minutes, 15X with pestle B. Samples were filtered through 30 um MACS strainers (Myltenyi, 130-098-458) and an additional 1 mL lysis buffer was added. Samples were centrifuged (500 rcf, 5 minutes, 4°C) and supernatant was discarded. Samples were resuspended in 750 uL wash buffer: 5mM MgCl­2, 10mM Tris buffer pH 8.0, 25 mM KCl, 1mM DTT, 27U/mL Protector RNase inhibitor, 1% bovine serum albumin. Samples were filtered through 30 um MACS strainers and centrifuged (500 rcf, 5 minutes, 4°C). Supernatant was discarded and pellets were resuspended in 1 mL wash buffer. Nuclei were stained with propidium iodide at a final concentration of 0.4ug/mL. Sorting was performed at the University of Michigan Flow Cytometry Core on a MoFlo Astrios Cell Sorter (Beckman Coulter). Sorted nuclei were collected in pre-washed tubes containing 100 uL wash buffer.
Library preparation and sequencing were performed by the University of Michigan Advanced Genomics Core using the 10X Genomics single-nucleus 3’ gene expression platform targeting 10,000 nuclei per sample and 60,000 reads per nucleus. Paired-end sequencing was performed on the NovaSeq6000.
 
Library strategy RNA-Seq
Library source transcriptomic single cell
Library selection cDNA
Instrument model Illumina NovaSeq 6000
 
Data processing Data preprocessing was performed by the University of Michigan Advanced Genomics Core using the 10X Genomics Cell Ranger pipelines (cellranger mkfastq, cellranger count, and cellranger multi).
Assembly: Mm10
Supplementary files format and content: Tab-separated values and matrix files
 
Submission date Jan 30, 2024
Last update date Jul 05, 2024
Contact name Sophie Hill
E-mail(s) sfhill@umich.edu
Organization name University of Michigan
Department Human Genetics
Lab Meisler Lab
Street address 1241 E. Catherine St
City Ann Arbor
State/province MI
ZIP/Postal code 48109
Country USA
 
Platform ID GPL24247
Series (1)
GSE254640 Long-term downregulation of SCN8A in mouse models of developmental and epileptic encephalopathy II
Relations
BioSample SAMN39678053
SRA SRX23455292

Supplementary file Size Download File type/resource
GSM8047578_6234_barcodes.tsv.gz 44.3 Kb (ftp)(http) TSV
GSM8047578_6234_features.tsv.gz 284.1 Kb (ftp)(http) TSV
GSM8047578_6234_matrix.mtx.gz 104.1 Mb (ftp)(http) MTX
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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