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| Status |
Public on Jun 30, 2020 |
| Title |
human_mfs_root |
| Sample type |
SRA |
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| Source name |
Aortic tissue
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| Organism |
Homo sapiens |
| Characteristics |
genotype: Marfan
Sex: male
anatomic segment: Aortic root
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
For aortic root/ascending samples, the aortic root was dissected completely and divided below the aortic valve. The ascending aorta was divided midway between the aortic valve and the takeoff of the brachiocephalic artery (Figure 2A). For descending thoracic aorta, the vessel was transected proximally just beyond the ligamentum arteriosum and distally at the diaphragm. Aortas were transferred to a separate dish and rinsed in PBS. Specimens were combined into groups with age and genotype-matched animals as biologic replicates. The aortas were then digested in enzymatic solution containing 2U/mL Liberase TM (Roche #05401127001) and 2U/mL elastase (Worthington #LS002279) in Hanks buffered saline solution (HBSS) for one hour. The cell suspension was filtered through a 70uM strainer, centrifuged and resuspended in cold HBSS. Suspensions were processed with FACS to discriminate live/dead cells and clumps/debris. Samples were then diluted and proprocessed using a 10x Genomics microfluidics chip to generate barcoded Gel Bead-In Emulsions (GEMs) according to manufacturer protocols
Individual sample libraries were generate from single cell GEM captures using the Chromium 3' version 3 library prep protocol (10X Genomics). QC was performed following cDNA generation and final library construction.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
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| Description |
Male (single patient)
g131_mfs_root_S1_L007
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| Data processing |
The sequenced data were processed into expression matrices with the ‘Cell Ranger Single cell software suite 3.1.0’. Raw base-call files from HiSeq4000 sequencer were demultiplexed to first generate FASTQ files using the cellranger mkfastq pipeline.
Reads were aligned to the mouse (mm10-3.0.0) or human (GRCh38-3.0.0) transcriptome, cell barcodes and unique molecular identifiers were filtered and corrected using the cellranger count pipeline
The final output filtered expression matrices were imported into the Seurat package in R and built into Seurat objects using the CreateSeuratObject function.
Standard quality control measures were then taken to exclude outliers representing doublets/clumps and free RNA. Cells with greater than 7.5% mitochondrial gene content were excluded
Data normalization, scaling and regression by mitochondrial content were then performed using the SCTransform command in Seurat.
Genome_build: mm10-3.0.0, GRCh38-3.0.0
Supplementary_files_format_and_content: Processed data files(.csv format) are exported expression matrices from the SCT assay within the Seurat package in R. Columns represent individual cell barcodes, rows represent genex and numerical values represent the normalized expression data.
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| Submission date |
Jun 29, 2020 |
| Last update date |
Jul 01, 2020 |
| Contact name |
Albert James Pedroza |
| E-mail(s) |
alpedroz@stanford.edu
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| Organization name |
Stanford University
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| Department |
Cardiothoracic Surgery
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| Lab |
Fischbein Lab
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| Street address |
870 Quarry Rd, Falk CVRB
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| City |
Stanford |
| State/province |
CALIFORNIA |
| ZIP/Postal code |
94304 |
| Country |
USA |
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| Platform ID |
GPL20301 |
| Series (1) |
| GSE153534 |
Aortic smooth muscle cell phenotype modulation during Marfan syndrome aneurysm formation |
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| Relations |
| BioSample |
SAMN15401978 |
| SRA |
SRX8636368 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4646673_human_root_expressionmatrix.csv.gz |
5.2 Mb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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