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| Status |
Public on Dec 21, 2021 |
| Title |
≥16N megakaryocytes from Pf4Cre-Srsf3fl/fl mice, replicate 2 |
| Sample type |
SRA |
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| Source name |
≥16N megakaryocytes
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| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
tissue: ≥16N megakaryocytes
genotype: Pf4Cre-Srsf3fl/fl
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| Extracted molecule |
total RNA |
| Extraction protocol |
Following fluorescence associated cell sorting based on CD41 and ploidy, low ploidy (8N only) and high ploidy (≥16N) megakaryocytes were pooled (52k±32k per sample) for RNA isolation using RNeasy Mini kit (Qiagen). RNA from washed platelets was isolated using TriReagent (Sigma-Aldrich).
Megakaryocyte RNA-seq libraries were prepared using SMARTer Stranded Total RNA Low Input Sample Prep kit (Clontech) and RNA-sequencing by Illumina HiSeq1500. Platelet RNA-seq libraries were prepared from 10ng of total RNA using an Ion AmpliSeq Transcriptome Mouse Gene Expression Kit and libraries were sequenced on an Ion Proton Sequencer (Thermo Fisher)
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 1500 |
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| Description |
Megakaryocytes_DEG
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| Data processing |
For megakaryocyte data, the raw read were trimmed using Trimmomatic with default parameters. The reads were aligned with STAR to Ensembl mm10 reference genome. The aligned files were deduplicated using the Picard MarkDuplicates function, ["Picard Toolkit." 2019 Broad Institute, Github Repository. http://broadinstitute.github.io/picard/; Broad Institute]. To obtain levels of gene expression, read counts were generated with the Subread function FeatureCounts. Genes with 1 count per million (CPM) in at least two samples were kept for further analysis. Normalisation and differential gene expression was conducted with EdgeR.
For platelet data, Torrent Suite Software v5.6 (Thermo Fisher) was used for read processing, base calling, alignment to the reference transcriptome (mm10) and gene expression counts. Differential gene expression analysis was performed using DESeq2.
Genome_build: mm10
Supplementary_files_format_and_content: .csv file containing log-fold change, p-value, FDR and read counts for DEGs
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| Submission date |
Aug 03, 2020 |
| Last update date |
Dec 21, 2021 |
| Contact name |
Minna-Liisa Anko |
| E-mail(s) |
minni.anko@monash.edu
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| Organization name |
Monash University
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| Department |
Department of Anatomy and Developmental Biology
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| Street address |
15 Innovation Walk
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| City |
Clayton |
| State/province |
VIC |
| ZIP/Postal code |
3800 |
| Country |
Australia |
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| Platform ID |
GPL18480 |
| Series (1) |
| GSE155620 |
RNA Binding Protein SRSF3 confers an essential role in megakaryocyte maturation and platelet production |
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| Relations |
| BioSample |
SAMN15711786 |
| SRA |
SRX8873884 |
| 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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