Hello!
I am conducting differential expression analysis on a subset of plant transcripts. I have decided to go with Salmon+tximport+DESeq2. I am using the pseudoalignment mode of salmon on fastp-trimmed fastq files. Salmon index is run with '-k=31' and quant is run with '--validateMappings' flags. I have run quant for the same samples twice and have observed significant variations in the TPM and NumReads. Between both the runs, about 82% of the transcripts were found to be common DETs by DESeq2.
I want to know if this margin of reproducibility is common or is there a way to increase the reproducibility of salmon quant?
Any and all suggestions are welcome!
Thank you
Can you say with confidence that all parameters, that means code, software versions, reference genome/transcriptome annotations were precisely the same? What is "significant variation"? Can you show some correlation plots to support this?