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Sample GSM7596915 Query DataSets for GSM7596915
Status Public on Oct 01, 2023
Title Rhabdomys_NeunPos1b2
Sample type SRA
 
Source name Retina
Organism Rhabdomys pumilio
Characteristics tissue: Retina
cell type: Retinal neurons
Treatment protocol None
Extracted molecule nuclear RNA
Extraction protocol Frozen retinal tissues were homogenized in a Dounce homogenizer in 1ml Tris-based lysis buffer with 0.1% NP-40. Nuclei were stained with NEUN/RBFOX3 and/or CHX10/VSX2, washed once, pelleted at 500g for 5min, resuspended in PBS/BSA and stained with DAPI. NEUN+ and/or CHX10+ DAPI+ single nuclei were collected using a flow cytometer. The sorted nuclei were pelleted again, resuspended in PBS/BSA solution, and adjusted to a concentration of 1000 nuclei/µL. ~12µL of the nuclei suspension was loaded into a 10X Chromium Single Cell Chip (10X Genomics, Pleasanton, CA) with a targeted recovery of 8000 nuclei.
Single nuclei libraries were generated using the Chromium 3’ V3.1 platform (10X Genomics, Pleasanton, CA) according to the manufacturer’s protocol. Briefly, single nuclei were partitioned into Gel-beads-in-EMulsion (GEMs) where nuclear lysis and barcoded reverse transcription of RNA would take place to yield full-length cDNA; this was followed by amplification, enzymatic fragmentation, and 5’ adaptor and sample index attachment to yield the final libraries.
Single nucleus RNA sequecning (10X Genomics)
 
Library strategy RNA-Seq
Library source transcriptomic single cell
Library selection cDNA
Instrument model Illumina NovaSeq 6000
 
Description Rhabdomys_count_mat.csv
Data processing We used cellranger (v7.0, 10X Genomics) to align the sc- and snRNA-seq datasets, following manufacturer’s instructions. For each species, sequencing reads were demultiplexed into distinct samples and the .fastq.gz files corresponding to each sample were aligned to reference transcriptomes to obtain binary alignment map (.bam) files.
Assembly: Rhabdomys_pumilio_2.0
Supplementary files format and content: To include both exonic and intronic reads in the quantification of gene expression for each sample, regardless of cellular or nuclear origin, we applied velocyto to the corresponding .bam files. This generated two separate gene expression matrices (GEMs; genes x cells) for each sample, corresponding to “spliced” and “unspliced” reads. The two GEMs were summed element by element to obtain “total” GEM for each sample.
Supplementary files format and content: GEMs from different sample runs were combined (column-wise concatenated) to yield a full species’ GEM.
 
Submission date Jul 12, 2023
Last update date Oct 01, 2023
Contact name Joshua William Hahn
E-mail(s) joshhahn@berkeley.edu
Organization name UC Berkeley
Department Chemical and Biomolecular Engineering
Lab Karthik Shekhar
Street address Stanley Hall
City Berkeley
State/province CA
ZIP/Postal code 94720
Country USA
 
Platform ID GPL33579
Series (2)
GSE237210 Evolution of neuronal cell classes and types in the vertebrate retina [Rhabdomys]
GSE237215 Evolution of neuronal cell classes and types in the vertebrate retina
Relations
BioSample SAMN36418862
SRA SRX20999104

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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