Kentaro Yoshida and Diane Saunders at TSL.
We used mRNAseq data from infected (AT1: data/ash_dieback/mixed_material/ashwellthorpe_AT1) and uninfected (ATU1: data/ash_dieback/fraxinus_excelsior/Ashwellthorpe_ATU1) to assess differential expression of Ash genes during infection by Chalara fraxinea.
- First, we aligned the reads from the infected sample (AT1) against the transcriptome assembly (assembled with the programme trinity) of an uninfected sample from the same location (ATU1).
- We aligned the reads from the uninfected sample against the assembly of this sample for comparison.
- Next, we counted the number of reads that mapped against unique positions in each transcript. Any reads that mapped to multiple locations were discarded.
- Finally, we ranked the counts based on their expression level (number of reads mapped). This is because the uninfected sample had many more reads than the infected sample so considering the total number of mapped reads would not be informative.
Usually we would run a normalisation step but the ranking should be sufficient to highlight any vast differences in the expression profiles. We would also like to run statistical analysis but so far we have only assessed one replicate and so this is not possible with the present data.