I have whole genome data in paired end fastq files. Before going for its alignment, I checked some of the samples for quality using FastQC but I'm getting strange results as seen in the attachments. Shall I move ahead with such data or need to trim these sequences or discard the data? I've majority of samples like this!




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Hi,
I would suggest you trim the leftmost part of the sequences. Given that you still have some high quality bases at the 3'-end, it will be good to set a Q-score threshold during trimming. By doing this you will end up with reads of varying lengths.
http://www.youtube.com/watch?v=bz93ReOv87Y
HTH
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Your Kmer plot suggests that a reasonable proportion of your library contains very short inserts (we can't really tell what proportion from the graph - you'd need the accompanying table for that).
Is it possible you've just made a library with extremely short inserts and that this is why the quality is dropping so quickly? Given the adapter contamination you probably need to adapter trim your data anyway for it to be of any use, so you might as well quality trim while you're at it.
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by GATTACATLove this - good data definitely starts from good input, and poor input can only give relatively poor data. I particularly like the mention of Nanodrop/absorbance based methods for quantification. It's such a toss up if you'll get an accurate reading or what amounts to a randomly generated number, and a lot of library/sequencing related issues can be traced back to poor quant.
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by SEQadmin2
I’m not a sequencing expert. I’m a purification scientist who uses NGS to evaluate workflows my group develops. With this perspective, we think about the sample first and the NGS workflow second. The sequencer is an exceptionally honest reporter, but it can only report on what you give it, so whether you get clean, interpretable data from an NGS workflow is largely determined before you begin.
Here are nine questions we think about, in roughly the order they matter, before...-
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