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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Apr 09, 2024 |
| Title |
CF1 NO |
| Sample type |
SRA |
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| Source name |
human bone marrow-derived mesenchymal stromal cells
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| Organism |
Homo sapiens |
| Characteristics |
condition: No osteoporosis
cell type: mesenchymal stromal cells
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| Growth protocol |
MSCs were isolated from the collected marrow as previously described in detail by Cassidy et al. (2020). Briefly, the collected marrow was centrifuged and the pellet resuspended in (PBS, Gibco via Biosciences, Dun Laoghaire, Ireland, 14190094). The number of mononuclear cells (MNCs) were quantified and the cells were plated in tissue culture flasks in expansion medium (α-MEM with GlutaMAX™, Gibco, 32561029) supplemented with 10% fetal bovine serum (FBS, HyClone via ThermoFisher, Ballycollen, Ireland SV30160.03), 1% antibiotic antimycotic solution (Merck, Dublin, Ireland, A5955), and 1 ng/mL recombinant human FGF-basic (Peprotech, London, UK, 100-18B)) at 37 °C with 5% CO2. Following the first passage (P0), the MSCs were lifted from the growth surface with 0.25% trypsin-Ethylenediaminetetraacetic acid (EDTA, Gibco, 25200056), evenly distributed in freezing medium (10% DMSO (Sigma, Arklow, Ireland, D2650), 90% FBS), and cryopreserved in liquid nitrogen.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the QIAGEN RNEasy Plus Mini Kit (Qiagen, Germany) and according to the manufacturer's protocol. RNA quality and quantity was then assessed with Nanodrop (Thermo Fisher, USA) and Agilent Bioanalyser 2100 with RNA 6000 Nano LabChip Kit (Agilent Technologies, USA).
Optimal RNA characteristics for sequencing were set by LC Sciences, USA (RNA amount ≥ 2μg, 260/280 >1.8, 260/230 >1.0, RIN value ≥ 7.0) who were contracted to perform sequencing, quality control, mapping and assembly of reads, and assessment of differential expression of genes and transcripts.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Firstly, Cutadapt and perl scripts in house were used to remove the reads that contained adaptor contamination, low quality bases and undetermined bases.
Then sequence quality was verified using FastQC (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/). We used HISAT2 to map reads to the genome .
The mapped reads of each sample were assembled using StringTie. Then, all transcriptomes were merged to reconstruct a comprehensive transcriptome using perl scripts and gffcompare.
After the final transcriptome was generated, StringTie and Ballgown was used to estimate the expression levels of all transcripts.
StringTie was used to perform expression level for mRNAs by calculating FPKM. The differentially expressed mRNAs were selected with log2 (fold change) >1 or log2 (fold change) <-1 and with statistical significance (p value < 0.05) by R package Ballgown.
Supplementary_files_format_and_content: Excel files include FPKM values for each Sample
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| Submission date |
Nov 24, 2021 |
| Last update date |
Apr 09, 2024 |
| Contact name |
Fearon C Cassidy |
| E-mail(s) |
fearon.cassidy@mu.ie
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| Organization name |
Maynooth University
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| Street address |
Kathleen Lonsdale Institute of Human Health
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| City |
Maynooth |
| ZIP/Postal code |
W23 F2H6 |
| Country |
Ireland |
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| Platform ID |
GPL11154 |
| Series (1) |
| GSE189524 |
Altered gene expression in multiple key pathways in MSCs from people with osteoporosis compared to people who do not have osteoporosis |
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| Relations |
| BioSample |
SAMN23431606 |
| Supplementary data files not provided |
| Raw data not provided for this record |
| Processed data are available on Series record |
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