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| Status |
Public on Nov 30, 2022 |
| Title |
514_nucleus_2 |
| Sample type |
SRA |
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| Source name |
Nuclei
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| Organism |
Mus musculus |
| Characteristics |
genotype: WT
gender: Female
age: 9-mo
tissue: hippocampus
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| Extracted molecule |
total RNA |
| Extraction protocol |
The frozen brain tissues were homogenized by Dounce Homogenizer, and the homogenate was fixed by incubating with 3% PFA at room temperature for 10 min. Hoechst-postive single nuclei and Hoechst-negative single synapses were sorted out by FANS.
Total-RNA based transcriptome library was generated with MATQ-Drop chemistry.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
Base-calling was performed with the standard Illumina NextSeq500 procedure.
The 3’ polyA tail of cDNA on Read 1 was trimmed with cutadapt v3.1, the Read 2 covering cell barcode sequences was assigned to a pre-defined barcode list.
Reads with successfully assigned cell barcodes were extracted with umi_tools extract (v1.0.1). Extracted Read 1 was mapped to the mm10 genome with STAR v2.5.3a, and the uniquely mapped reads with mapping q score no smaller than 250 was used for downstream analysis.
The filtered reads were assigned to genes by featureCounts v2.0.1 with appropriate Gencode annotation gtf files. (stradness parameter: -s 2)
For the reads with unambiguously assigned gene features, umi_tools “count” command was used to generate the transcript-based or exon-based digital gene expression matrix (parameter: --per-gene --gene-tag=XT --per-cell –method=directional).
Nuclei with mitochondrial UMI fraction higher than 5% were removed. Synapses with mitochondrial UMI fraction smaller than 5% were removed. Next, genes on chrMT and rRNA genes (based on Ensembl annotation) were removed from downstream analysis.
Assembly: mm10
Supplementary files format and content: tab-delimited files include the transcript-based, intron-based, and exon-based UMI count matrix for each sample.
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| Submission date |
Mar 24, 2022 |
| Last update date |
Nov 30, 2022 |
| Contact name |
Chenghang Zong |
| E-mail(s) |
[email protected]
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| Organization name |
Baylor College of Medicine
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| Lab |
Zong Lab
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| Street address |
1 Baylor Plaza
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| City |
Houston |
| State/province |
TX |
| ZIP/Postal code |
77030 |
| Country |
USA |
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| Platform ID |
GPL19057 |
| Series (2) |
| GSE199344 |
High-Throughput Transcriptome Profiling of Single Nuclei and Single Synapses Using Single-Cell Total-RNA-Seq [Mm] |
| GSE199346 |
High-Throughput Transcriptome Profiling of Single Nuclei and Single Synapses Using Single-Cell Total-RNA-Seq |
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| Relations |
| BioSample |
SAMN26931464 |
| SRA |
SRX14598340 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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