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Sample GSM8522773

Query DataSets for GSM8522773

Status

Public on Oct 30, 2024

Title

Mouse, female sample1, sublibrary1

Sample type

SRA

Source name

Ventral pallidum, bilateral, pooled from 5 mice

Organism

Mus musculus

Characteristics

tissue: Ventral pallidum, bilateral, pooled from 5 mice
genotype: C57Bl6J

Extracted molecule

polyA RNA

Extraction protocol

Nuclear extraction for both mice and primates was performed with density gradient centrifugation according to the protocol described previously with modifications to scale down the reaction volume (85). Samples were moved from -80 ̊C and thawed on ice for 2 minutes. Tissues were transferred to a dounce homogenizer (Millipore Sigma, D8938-1SET) in homogenization buffer (0.25 M sucrose, 25 mM KCl, 5 mM MgCl2, 10 mM Tris-HCl, pH 8.0, 5 µg/mL actinomycin, 1% BSA, and 0.08 U/µl RNase inhibitor, 0.01% NP40) on ice. Samples were homogenized for 10 strokes with the loose pestle in a total volume of 1 mL, followed by 10 additional strokes with the tight pestle. The tissue homogenate was then passed through a 50 µm filter and diluted 1:1 with working solution (50% iodixanol, 25 mM KCl, 5 mM MgCl2, and 10 mM Tris-HCl, pH 8.0). Nuclei were layered onto an iodixanol gradient after homogenization and ultracentrifuged as described previously. After ultracentrifugation, nuclei were collected between the 30 and 40% iodixanol layers and diluted with resuspension buffer (1xPBS with 1% BSA, and 0.08 U/µl RNase inhibitor). Nuclei were centrifuged at 500 g for 10 min at 4°C and resuspended in resuspension buffer with 5 ng/µl of 7-AAD. FACS was carried out to further remove cellular debris. 7-AAD+ events were collected using a 100 µm nozzle on a BD FACSAria III instrument into a 1.5 mL microcentrifuge tube.
Nuclei were processed according to the manufacturer’s manual of 10X Genomics Chromium Single Cell Gene Expression 3’ V3.1 Assay.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NovaSeq 6000

Data processing

Raw sequencing data from individual libraries were processed and mapped to the reference genome and the gene-cell counts matrices from all snRNA-seq libraries of the same species were concatenated using 10X Genomics cellranger-v6.1.2 or cellranger-v7.0.1
Assembly: mouse: mm10, baboon: Panubis1.0, and macaque: Mmul_10
Supplementary files format and content: Gene matrix.rds contains gene-cell counts for all libraries, metadata.csv contains meta data for all libraries

Submission date

Sep 18, 2024

Last update date

Oct 30, 2024

Contact name

Lite Yang

E-mail(s)

yanglite1211@gmail.com

Organization name

Washington University in St. Louis

Street address

4922 Parkview Place