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per tile sequence quality decreases after QC with sickle

I'm trying to QC my illumina miseq paired end reads with sickle pe. but, as visualized in Fastqc per tile sequence quality, it seems that I can not get rid of low quality sequences from special tiles (2118) but somehow it even get worse by increasing quality threshold.

this is the screen-shot from my reads per tile sequence quality after trimming with default quality threshold of sickle pe (q 20):

https://postimg.org/image/s8igtzzc5/

and this is what I get after increasing that to q=30:

https://postimg.org/image/q30kiflbr/

any Idea about what happens around 2118?

sickle fastqc per tile sequence quality
• 2,943 views • link
written by mary • mary

Instead of dropbox links consider posting the images on imgur.com (or other free image hosting sites) and including the links in your post (use the ctrl+G option when editing the post).

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written by GenoMax GenoMax

1 answer

I suspect it turns red if 100% of the reads are gone from a tile, in which case the calculated average quality would be 0 to avoid a division by zero problem.

For this kind of issue, I suggest using FilterByTile rather than blanket-applying extremely high quality-trimming thresholds like Q30 to your entire dataset, which incurs bias and can greatly damage the utility of the data for many purposes.

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written by Brian Bushnell Brian Bushnell

seems reasonable! thanks for your help!

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written by mary • mary

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