2024-2: 7d-Homework
- Vicki Thomson
Exercises for you to try:
Redo the R analysis for the human drosha dataset
Precomputed results from session 6:
We ran the small RNA seq samples against the MirGeneDB database and the results can be found at:
/work/training/2024/smallRNAseq/runs/run4_mirBase/results/mirna_quant/edger_qc/mature_counts.csv
/work/training/2024/smallRNAseq/data/human_disease/metadata_microRNA.txtLet’s create a “DESeq2” folder and copy the files needed for the statistical analysis:
cp $HOME/workshop/2024-2/session6_smallRNAseq/scripts/transpose_csv.py $HOME/workshop/2024-2/session6_smallRNAseq/runs/run3_drosha_miRBase/DESeq2
cp $HOME/workshop/2024-2/session6_smallRNAseq/data/metadata_microRNA.txt $HOME/workshop/2024-2/session6_smallRNAseq/runs/run3_drosha_miRBase/DESeq2
cp /work/training/2024/smallRNAseq/runs/run3_MirGeneDB/results/mirna_quant/edger_qc/mature_counts.csv $HOME/workshop/2024-2/session6_smallRNAseq/runs/run3_drosha_miRBase/DESeq2
cd $HOME/workshop/2024-2/session6_smallRNAseq/runs/run3_drosha_miRBase/DESeq2To transpose the initial “mature_counts.csv” file do the following:
python transpose_csv.py --input mature_counts.csv --out mature_counts.txtDifferential expression analysis using RStudio
Run analysis script in RStudio
Pre-steps: Open RStudio, Create a new R script ('File'->'New File'-> ‘R script’), Hit the save button and save this file in the working directory you created above (H:\workshop\2024-2\session6_smallRNAseq\runs\run3_drosha_miRBase\DESeq2). Name the R script ‘DESeq2.R’.
Step 1: LOAD PACKAGES
Step 2: IMPORT DATA
Step 3: LOOKING FOR OUTLIERS AND BATCH EFFECTS - TRANSFORM DATA
Step 4: LOOKING FOR OUTLIERS AND BATCH EFFECTS - VISUALISE DATA (PCA)
Step 5: LOOKING FOR OUTLIERS AND BATCH EFFECTS - VISUALISE DATA (HEATMAP)
Step 6: LOOKING FOR DIFFERENTIALLY EXPRESSED GENES
Step 7: LOOKING FOR DIFFERENTIALLY EXPRESSED GENES - VISUALISATION (VOLCANO PLOT)
Step 8: REMOVE OUTLIERS and LOOKING FOR DIFFERENTIALLY EXPRESSED GENES - VISUALISATION (PCA, VOLCANO PLOT)
Step 9: DIFFERENTIAL EXPRESSION ANALYSIS - HEATMAP and DENDROGRAM