I am currently analysing RNA-SEQ data (differential expression). I have 20 samples in total: 5 conditions in 2 donors and each donors- each donor has 2 repeats (cell type- human bronchial epithelial cells).
I have 24fastq files per sample- these have been QC'd in fastqc and run through tophat to produce 1 BAM file per sample.
The BAM files were then run through cufflinks for transcript assembly (gtf files).
I have merged all gtf files in cuffmerge and am not sure whether to use cuffdiff or cuffnorm.
Could i ask if the pipeline i have used so far is correct and also ask for advice how to calculate differentail expression.