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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Feb 21, 2023 |
| Title |
NeuroD1, scRNA |
| Sample type |
SRA |
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| Source name |
gut
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| Organism |
Mus musculus |
| Characteristics |
tissue: gut
genotype: NeuroD1-Cre;tdTom
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| Extracted molecule |
total RNA |
| Extraction protocol |
Intestinal tissue was obtained from Neurog3-Cre; lsl-tdTomato mice (1 adult male), or Neurod1-Cre; lsl-tdTomato (3 adult females), cut longitudinally, washed (cold phosphate-buffered saline or PBS), cut into small ~5 mm pieces, and incubated (gentle agitation, 20 minutes, 4C) in EDTA solution (20 mM EDTA-PBS, Ca/Mg-free) in LoBind Protein tubes (Eppendorf 0030122216). The specimen was shaken, the tissue allowed to settle, and the supernatants collected. The residual tissue was again incubated similarly with EDTA solution, and supernatants were combined, and centrifuged (300 g, 5 minutes, 4C) Pellets were washed (2x, PBS (Ca/Mg-free) supplemented with 5% Fetal Bovine Serum (FBS), 4C) and incubated (37C, 2 minutes) in protease solution (TrypLE express, Thermo Fisher 12604013) supplemented with DNase (100 U/ml, Worthington Biochemical LK003172). The suspension containing dissociated cells was centrifuged (300 g, 5 minutes), washed (2x, PBS (Ca/Mg-free) containing 5% FBS, 4C) The resulting pellet was resuspended in FACS buffer (5% FBS in DMEM/F12, HEPES, no phenol red) containing DNase (100 U/ml), TO-PRO-3 (Thermo Fisher T3605, 1:10,000) to label dead cells, and Calcein Violet (Thermo Fisher 65-0854-39, 1:10,000) to label living cells. Cells were filtered (1 x 70-um, 1 x 40-um) and tdTomato+, Calcein Violet+, TO-PRO-3- cells were collected by fluorescence activated cell sorting using a FACS Aria (BD Biosciences).
Collected cells were loaded into the 10x Genomics Chromium Controller, and cDNA prepared and amplified according to manufacturer’s protocol (10x Genomics, Chromium single cell 3’ reagent kit v3, 12 cycles per amplification step).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
10x Genomics
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| Data processing |
Sequence reads were aligned to the mm39 mouse transcriptome reference, and feature barcode matrices were generated using 10x Genomics CellRanger.
Assembly: mm39
Supplementary files format and content: HDF5 Feature-Barcode Matrix from CellRanger
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| Submission date |
Feb 01, 2023 |
| Last update date |
Feb 21, 2023 |
| Contact name |
Judith Kaye |
| Organization name |
Harvard University
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| Department |
Cell Biology
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| Lab |
Liberles
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| Street address |
240 Longwood Ave, LHRRB 6th floor
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| City |
Boston |
| State/province |
Massachusetts |
| ZIP/Postal code |
02115 |
| Country |
USA |
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| Platform ID |
GPL19057 |
| Series (1) |
| GSE224223 |
Enteroendocrine cell lineages that differentially control feeding and gut motility |
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| Relations |
| BioSample |
SAMN33005428 |
| SRA |
SRX19244979 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7017776_NeuroD1_filtered_feature_bc_matrix.h5 |
15.7 Mb |
(ftp)(http) |
H5 |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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