FastQC Per sequence GC count always fails after trimmomatic?
0
0
Entering edit mode
3.9 years ago
DNAngel ▴ 250

HI all,

Just a quick question on theory I suppose. After running trimmomatic on my fastq files with the following, generally used parameters such as:

ILLUMINACLIP:adapters:TruSeq2-PE.fa:2:30:10 
SLIDINGWINDOW:4:20
HEADCROP:5
MINLEN:36

And then running FastQC on the trimmed, paired dataset - most quality checks seem to be okay except the Per sequence GC content goes from being okay (untrimmed dataset) to failing it. Now, I think it is because after trimming poor quality bases and such I do get a variety of sequence lengths from 36-146 and I believe this is driving the sudden increase in GC content in especially short sequences.

My question here is, is this correct? Is this why it would fail GC content and I can safely ignore this warning? It makes sense to me that shorter sequences would suddenly have high %GC content and this is causing this quality check to fail but I don't know if this is actually something we expect.

Thank you!

EDIT:

Image to show the FastQC results for two samples - it looks like my trimming does not affect GC content at all.

https://i.ibb.co/FKDsWQT/fastqc-results.jpg

enter image description here EDIT2:

I should mention this is whole genome sequence data not RNAseq data.

fastqc trimmomatic • 2.0k views
ADD COMMENT
0
Entering edit mode

Is your organism expected to have a GC rich genome? There is no change essentially after the trimming.

ADD REPLY
0
Entering edit mode

I am not sure actually - first time working with Hymenoptera species. Some quick research says there are GC-rich domains but I think most eukaryotes would have some regions of the genome be more GC rich than others for different purposes.

ADD REPLY
0
Entering edit mode

If you don't have a concrete reason to think that this observation is problematic then you can move ahead with rest of your analysis.

ADD REPLY
0
Entering edit mode

Okay thanks for your feedback!

ADD REPLY

Login before adding your answer.

Traffic: 1312 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6