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| Status |
Public on Jan 17, 2024 |
| Title |
day15,treat12,scRNAseq |
| Sample type |
SRA |
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| Source name |
skin
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| Organism |
Mus musculus |
| Characteristics |
tissue: skin
cell type: skin cells
genotype: db/db(C57BLKS/J)
treatment: TSeL with light irradiation (TSeL+L)
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| Extracted molecule |
total RNA |
| Extraction protocol |
The skin tissues without separating dermis and epidermis were dissociated from the diabetic mice wound and washed with pre-cold PBS buffer. And skin tissues were minced as smaller as possible in digestive enzyme mixture and then incubated at 37°C for 1.5 h. Digestive enzyme mixture contains collagenase IV ( 200 mg/ml), Deoxyribonuclease I (1 mg/ml) , HEPES buffer solution (25mM) and RPIM-1640 medium supplemented with 10% fetal bovine serum. Subsequently, digestion process was terminated by adding the medium, and the digested skin tissues were further milled and filtered through a 40 μm strainer. Cell count and viability was estimated using fluorescence Cell Analyzer (Countstar® Rigel S2) with AO/PI reagent after removal erythrocytes (Miltenyi 130-094-183). Afterward, fresh cells were washed with ice-cold RPMI 1640 containing 2% FBS for twice and then resuspended at 1×106 cells per ml in 1×PBS and 0.04% BSA.
Single-cell RNA-seq libraries were prepared using SeekOne® Digital Droplet (SeekOne® DD) Single Cell 3’ library preparation kit V2.1. Briefly, appropriate number of cells were mixed with reverse transcription reagent and then loaded to the sample well in SeekOne® DD Chip S3 (ChipS3). Subsequently, Barcoded Hydrogel Beads (BHBs) and partitioning Oil were dispensed into corresponding wells separately in chip. After emulsion droplet generation reverse transcription were performed at 42°C for 90 minutes and inactivated at 85°C for 5 minutes. Next, cDNA was purified from broken droplet and amplified in PCR reaction. The amplified cDNA product was then cleaned, fragmented, end repaired, A-tailed and ligated to sequencing adaptor. Finally the indexed PCR were performed to amplified the DNA representing 3’ polyA part of expressing genes which also contained Cell Bar code and Unique Molecular Index. The indexed sequencing libraries were cleanup with SPRI beads, quantified by quantitative PCR (KAPA Biosystems KK4824) and then sequenced on illumina NovaSeq 6000 with PE150 read length
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
SeekSoul Tools pipeline were used to process the cleaned reads and generated the transcript expression matrix.Firstly, the cell barcodes and UMI sequences were extracted based on the defined pattern about the localization of the barcode, linker and UMI within a read. The barcode was corrected with whitelist. The corrected barcode , together with UMI, were put in the header of their corresponding reads.Secondly, the reads were mapped to the reference genomes using STAR 2.7.2a.Then, the reads with barcode and UMI information were assigned to transcriptome using featureCounts of package Subread v2.0.1.
Assembly: mm10
Supplementary files format and content: matrix files
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| Submission date |
Jan 12, 2024 |
| Last update date |
Jan 17, 2024 |
| Contact name |
Chunqiu Zhang |
| E-mail(s) |
zhangcq@nankai.edu.cn
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| Phone |
15101068459
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| Organization name |
Nankai University
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| Street address |
NO 94 Weijin Road
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| City |
Tianjin |
| ZIP/Postal code |
300071 |
| Country |
China |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE253098 |
Gene expression profile at single cell level of skin cells isolated from db/db mice |
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| Relations |
| BioSample |
SAMN39422820 |
| SRA |
SRX23203325 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8013670_Treat12_barcodes.tsv.gz |
88.2 Kb |
(ftp)(http) |
TSV |
| GSM8013670_Treat12_features.tsv.gz |
254.0 Kb |
(ftp)(http) |
TSV |
| GSM8013670_Treat12_matrix.mtx.gz |
91.6 Mb |
(ftp)(http) |
MTX |
SRA Run Selector |
| Raw data are available in SRA |
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