ViReMa not working after adapter trimming

Question

[berelly]

Hi, I have a viral RNA-seq dataset that I am trying to run through ViReMa to look for deletion junctions. When I input my fastq files directly into ViReMa without trimming adapters first, my results look about how I would expect (multiple junctions returned with several reads each). However, when I trim adapters with cutadapt prior to running ViReMa, I get very few junctions returned, or (usually) no junctions at all.

Is it possible that adapter contamination would be causing artifactual junctions to be called? Or am I doing something super wrong in my trimming step and somehow removing information that ViReMa needs to proceed?

My cutadapt parameters look like this (just trimming Illumina universal adapters):

cutadapt -a AGATCGGAAGAG -o sample_cutadapt.fastq sample.fastq

And my ViReMa parameters looks like this:

python ViReMa.py --MicroInDel_Length 20 -DeDup --Defuzz 3 --Seed 25 --N 1 --X 2 --Output_Tag sample -ReadNamesEntry --p 6 bowtie1index/virus sample_cutadapt.fastq sample.results 

I'm new to this type of analysis, so any help is appreciated.

Answer 1

[berelly]

The problem was over-trimming, just needed to add an "X" to the end of my adapter sequence in the cutadapt command so that adapters were only trimmed from the 3' end (not from within anywhere in the sequence).

ie:

cutadapt -a AGATCGGAAGAGX -o sample_cutadapt.fastq sample.fastq