edgeR practice: crab 2019 data
Haven’t posted in a long long time… so in today’s post I’ll detail what I’ve been up to with the crab project: paper progress and trying out edgeR for differential gene expression analyses.
edgeR in the Ecology of Infectious Marine Disease course at FHL in summer 2019 to find differentially expressed genes in eelgrass with and without exposure to a pathogen, Labyrinthula zostera. Very convenient timing that I took that class becuase it was before any analyses were being performed for the crab project.
I went back thorugh the github repo we had from the class and tested out
edgeR by reproducing this script: https://github.com/eimd-2019/project-EWD-transcriptomics/blob/master/analyses/EdgeR/GRASS_START_GENE_MEE_Contrasts_CAB_7_10_nonZostera_at.R
Things went smoothly and I was able to reproduce everything after I got the ‘targets.txt’ file from Colleen.
Trying with crab data: 2019 dataset
The 2019 RNAseq data was a combination of temperature treatments, but there were four main groupings:
- Day 26 Uninfected
- Day 26 Infected
- Day 12 Uninfected
- Day 12 Infected
Steven and Sam helped with getting a file of gene counts for each sample (link: here)
And I made a ‘targets’ file that lists out the samples and their treatments (link: targets_2019.txt). I originally had the sample treatments listed as “26_Uninf”, etc., but there’s a part in the script where contrasts between different groupings need to be made, and having numbers in the treatment descriptions was messing things up, so I changed it to:
- “E” = day 12 (stands for ‘earlier’)
- “L” = day 26 (stands for ‘later’)
- “U” = uninfected
- “I” = infected
After that was fixed, the code worked well.
Link to the Rmd file of testing out
edgeR with the crab 2019 dataset:
It worked, but I need to go through more slowly and with the manual at hand to make sure all the steps make sense for the crab data. (Manual for
Crab project updates
Paper draft: here
- have written methods for RNA extraction for each data set
- have written what I can for methods of crab collection and experimental set-up
- starting to get some differential gene expression progress in that I’m testing out
edgeRand working on comparisons
- will work on making a mortality figure
Other: We got the data back from NWGC that I sent out a while back (Sam’s working with it) - notebook post detailing what the samples are: here
This week (4/13) - focusing on crab results
Next week (4/20) - working on discussion
week after (4/27) - working on intro
Goal to have draft of full thesis done by somewhere around May 4th to send to committee.