In order to maintain reproducibility when carrying out the downstream analysis, we have developed a custom Nextflow pipeline which integrates the alignment outputs into a single workflow.
This workflow makes use of a custom Docker container which contains all required packages.
If you would like to re-run the entire downstream analysis, you will need to first align the data by following the steps in the Quick start guide.
After aligning the data, you will need to create a samplesheet csv containing the paths to all of the alignment output folders. You can find an example of this here.
The entire downstream analysis can then be carried out by executing using the following shell script.
export NXF_VER=20.07.1
nextflow run ./NF-downstream_analysis/main.nf \
-c <PATH_TO_CONFIG> \
--input <PATH_TO_DOWNSTREAM_ANALYSIS_SAMPLESHEET.CSV> \
--outdir output/NF-downstream_analysis \
-resume
If do not want to re-run the alignment, but would like to run the downstream analysis from the count files, you can run RStudio server from within the Docker container. This will ensure that you have all of the same packages and dependencies required to carry out the analysis.
RNAseq read counts (Lmx1aE1 vs Sox3U3, SOX8OE and SmartSeq2) and ChIP/ATAC peaks files are embedded in the repository - they will be downloaded automatically when the repository is cloned.
To interactively explore the data, follow these steps:
git clone --recurse-submodules https://github.com/alexthiery/otic-reprogrammingdocker pull alexthiery/otic-reprogramming-r_analysis:latestdocker run --rm -e PASSWORD=password -p 8787:8787 -v <PATH_TO_LOCAL_REPOSITORY>:/home/rstudio alexthiery/otic-reprogramming-r_analysis:latestlocalhost:8787 in the browser to open RStudiorstudio and the password passwordFiles tab in R studio by following the path ./NF-downstream_analysis/bin/<FILE_OF_INTEREST>