generated from statOmics/Rmd-website
-
Notifications
You must be signed in to change notification settings - Fork 3
Commit
This commit does not belong to any branch on this repository, and may belong to a fork outside of the repository.
- Loading branch information
1 parent
42e55e7
commit 4f9b81d
Showing
4 changed files
with
52 additions
and
0 deletions.
There are no files selected for viewing
Binary file not shown.
This file contains bidirectional Unicode text that may be interpreted or compiled differently than what appears below. To review, open the file in an editor that reveals hidden Unicode characters.
Learn more about bidirectional Unicode characters
Original file line number | Diff line number | Diff line change |
---|---|---|
@@ -0,0 +1,50 @@ | ||
--- | ||
title: Differential transcript usage analysis | ||
author: "Lieven Clement" | ||
date: "[statOmics](https://statomics.github.io), Ghent University" | ||
output: | ||
html_document: | ||
theme: flatly | ||
code_download: false | ||
toc: false | ||
toc_float: false | ||
number_sections: false | ||
bibliography: msqrob2.bib | ||
|
||
--- | ||
|
||
<a rel="license" href="https://creativecommons.org/licenses/by-nc-sa/4.0"><img alt="Creative Commons License" style="border-width:0" src="https://i.creativecommons.org/l/by-nc-sa/4.0/88x31.png" /></a> | ||
|
||
### satuRn vignette | ||
|
||
We developed satuRn for fast differential transcript usage analysis on bulk and single cell transcriptomics data. | ||
Read and run the vignette of satuRn to learn howto work with satuRn | ||
|
||
![satuRn vignette](https://www.bioconductor.org/packages/release/bioc/vignettes/satuRn/inst/doc/Vignette.html) | ||
|
||
### Airway example | ||
|
||
The data used in this workflow comes from an RNA-seq experiment where airway smooth muscle cells were treated with dexamethasone, a synthetic glucocorticoid steroid with anti-inflammatory effects (Himes et al. 2014). Glucocorticoids are used, for example, by people with asthma to reduce inflammation of the airways. In the experiment, four human airway smooth muscle cell lines were treated with 1 micromolar dexamethasone for 18 hours. For each of the four cell lines, we have a treated and an untreated sample. For more description of the experiment see the article, PubMed entry [PMID: 24926665](https://pubmed.ncbi.nlm.nih.gov/24926665/), and for raw data see the GEO entry [GSE52778](https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE52778). | ||
|
||
We will conduct a DTU analysis by starting from quantification using the fast alignment-free transcript mapper Salmon. | ||
|
||
This can be done as follows: | ||
|
||
- Building index with salmon | ||
``` | ||
salmon index --gencode -t gencode.v32.transcripts.fa -i gencode.v32_salmon_index | ||
``` | ||
|
||
- Mapping one sample with salmon: | ||
``` | ||
salmon quant -i gencode.v32_salmon_index -l A --gcBias -1 SRR1039508_subset_1.fastq -2 SRR1039508_subset_2.fastq --validateMappings -o quant/SRR1039508_subset_quant | ||
``` | ||
|
||
More details can be found in [Intro to salmon](https://combine-lab.github.io/salmon/getting_started/). | ||
The mapped output from Salmon can be found at: [data](https://github.com/statOmics/SGA/archive/airwaySeqData.zip). | ||
|
||
1. Import the transcript level counts with the tximeta package: see [tximeta](https://bioconductor.org/packages/release/bioc/vignettes/tximeta/inst/doc/tximeta.html#Running_tximeta) vignette on bioconductor | ||
|
||
2. Run a DTU analysis on the transcript level counts with satuRn | ||
|
||
Tip: I had to use the argument `importer=read.delim` in the tximeta function because the default function to read the files returned an error on my laptop. |
This file contains bidirectional Unicode text that may be interpreted or compiled differently than what appears below. To review, open the file in an editor that reveals hidden Unicode characters.
Learn more about bidirectional Unicode characters