|
| Status |
Public on Mar 24, 2024 |
| Title |
Gut epithelial cells sham vs pMCAO 24 h (sham biol rep 2) [KP_60_S21] |
| Sample type |
SRA |
| |
|
| Source name |
Gut epithelial cells sham vs pMCAO 24 h (sham)
|
| Organism |
Mus musculus |
| Characteristics |
cell type: Small intestine epithelial cell
strain: C57BL6/J
treatment: Sham
time: 24 h
|
| Treatment protocol |
Sham or pMCAO surgery for 24 hours
Epithelial cells from the whole small intestines were isolated via FACS
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was harvested using Rneasy mini plus kit (Qiagen). extracted RNA was tested using an RNA 6000 Pico kit on the Agilent 2100 Bioanalyser, 2.0 ng of total RNA was used for the construction of sequencing libraries.
An RNA cleavage approach was carried out to remove ribosomal RNA. Library construction and cDNA synthesis were performed using the Trio RNA-sequencing library preparation kit (protocol M01440v2, 2017; Tecan). Manual denaturing and on-board clustering were carried out using 70pM of library pool containing 1% PhiX (protocol 1000000109376 v3, 2020; Illumina). Sequencing was carried out on the NextSeq 2000 (Illumina) with P3 reagents and paired-end reading of 100 base pairs. Approximately 35 million reads were sequenced for each sample
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
NextSeq 2000 |
| |
|
| Data processing |
Raw fastq files have been analysed with RNAsik pipeline (Tsyganov et al. 2018) to produce raw genes count matrix and various quality control metrics.
For this analysis RNAsik pipeline (Tsyganov et al. 2018) ran with STAR aligner option (Dobin et al., 2013) mapping raw reads to the mus musculus GRCm38-mm10 (GCF_000001635.20, NCBI) reference genome, reads were quantified with featureCounts (Liao, Smyth, and Shi 2014).
Raw counts were then analysed with Degust (Powell 2015) web tool to do differential expression analysis to produce list of differentially expressed genes and several quality plots including classical multidimensional scaling (MDS) and MA plots
In this analysis limma voom (Law et al. 2014) was used for differential expression analysis. Degust (Powell 2015) largely follows limma voom workflow with typical counts per million (CPM) library size normalisation and trimmed mean of M values (TMM) normalisation (Robinson and Oshlack 2010) for RNA composition normalisation.
Assembly: GRCm38-mm10 (GCF_000001635.20, NCBI)
Supplementary files format and content: raw gene counts, text file, tab separated, includes all samples
|
| |
|
| Submission date |
Oct 26, 2022 |
| Last update date |
Mar 24, 2024 |
| Contact name |
Deanna S Deveson Lucas |
| Organization name |
Monash University
|
| Lab |
Monash Bioinformatics Platform
|
| Street address |
15 Innovation walk, Monash University, Clayton
|
| City |
Melbourne |
| State/province |
Victoria |
| ZIP/Postal code |
3800 |
| Country |
Australia |
| |
|
| Platform ID |
GPL30172 |
| Series (2) |
| GSE216581 |
Effect of stroke on the transcriptome of epithelial cells in the small intestines |
| GSE216972 |
Effect of stroke on the transcriptome in the small intestines |
|
| Relations |
| BioSample |
SAMN31455787 |
| SRA |
SRX18039509 |
