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FastQC Kmer Content

Hello, I am attempting to prepare some 150 PE Illumina data for genome assembly. I saw that my raw data still had universal adapters. In addition it also failed the kmer content section so I processed it with trimmomatic (version 0.39) with the following parameters:

ILLUMINACLIP:TruSeq3-PE-2.fa:2:30:10 LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:36

A second FastQC (version 0.11.5) of the file for forward paired reads shows that the adapters are removed, but the kmer content still fails. Specifically in the 135-139 nt region there is a drastic increase in certain kmers. I've shared an image of what I am seeing. Does anyone have suggestions for how to modify my trimmomatic parameters to correct this issue?

enter image description here

qc sequencing
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written by KSS • KSS

failed some test in FastQC does not mean that you have to do something with that part or that you can't proceed to next step.

Because of the confusion caused by k-mer module it is now disabled by default (from v.0.11.6 onwards). You can move forward with rest of analysis.

This is what release notes say: There is one major change which is that by default we now disable the kmer module. With the inclusion of the adapter plot the value of the information in the Kmer plot is often not great, and it is easy to confound it if there are any over-represented sequences, or primer compositional bias. Overall therefore we consider it best to not routinely include this module.

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written by GenoMax GenoMax

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