• All data for this analysis can be obtained from GEO via the study accession number GSE178187. The processed data can also be found in this repository under the folder data/aligned.
• Data from the Koenecke et al. paper can be found on GEO with the accession number GSE68983. The data was downloaded using the prefetch command from the SRA Toolkit. The aligned and quantified data can also be found in this repository under the folder data/koenecke_rnaseq.
• Annotation file used: Drosophila_melanogaster.BDGP6.22.97.chr.gtf
• Reference fasta file used: dm6.fa (Ensembl v97)
• The dorsal target genes were obtained from Stathopoulos et al.

Ensure that the annotation file is in the data/annotation folder before running the analysis

The analysis was carried out to compare gene expression in the wild type samples (WT_4, WT_5, WT_6) and the mutant loss-of-function samples (S926glc_4, S926glc5, S926glc_6), using the QC.Rmd, DE.Rmd, and V_E.Rmd scripts for quality control, differential expression, and visualisation and enrichment respectively.

If running from scratch, these scripts can be found in the analysis folder and need to be run in the sequential order of QC.Rmd > DE.Rmd > V_E.Rmd as each script generates files required by the subsequent scripts. If not running from scratch, the outputs of each script can be found in their respective output subfolder in the analysis main folder. If the repository is cloned as is, and the .Rproj file is used, there should have no issues running the scripts.

The same folder structure applies to the subanalyses.

Here we looked at the RNA-seq data from Koenecke et al.. The processed data can be found in data/koenecke_rnaseq. The data was aligned using STAR and RSEM (see Versions/Koenecke Analysis for versions) via a Snakefile, which can be found in the data/koenecke_rnaseq folder. Again, the same three scripts were used, and in the same order -- QC.Rmd, DE.Rmd, and V_E.Rmd.

4.2.46(2)-release x86_64-redhat-linux-gnu

## R version 4.0.5 (2021-03-31)
## Platform: x86_64-apple-darwin17.0 (64-bit)
## Running under: macOS Big Sur 10.16
## 
## Matrix products: default
## BLAS:   /Library/Frameworks/R.framework/Versions/4.0/Resources/lib/libRblas.dylib
## LAPACK: /Library/Frameworks/R.framework/Versions/4.0/Resources/lib/libRlapack.dylib
## 
## locale:
## [1] en_US.UTF-8/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8
## 

• STAR version: 2.7.1a
• RSEM version: v1.3.1

• STAR version: 2.7.9a
• RSEM version: v1.3.1

The versions of R packages used in the analysis can be found in the Session Info section of each .html file.