qseq files pattern of reads with no sequence

brentp · 08-26-2010, 09:27 AM

i dont believe this is a problem, just something i'm curious about:

normally we get our data from a sequencing center that returns a fastq file. this time, we have a bunch of qseq files like: s_3_1_0001_qseq.txt
in all of them, the first and last few hundred reads in the file have no sequence (just .'s for the length of the read). as one moves down from the top of the file (or up from the bottom), it starts that sequence appears on either end of the read, so it will look like:

Code:

G.TCTG..............................................................ACCCGGA.

(and the quality is all B's). going to the center of the file, the reads are all sequence.
i'm curious as to why this occurs at the ends of the file. is there some correspondence between position in the file and location on the cell.

thanks.

kmcarr · 08-26-2010, 09:44 AM

Your guess is correct. Reads are sorted by their x-coordinate (but apparently there's no sorting by y-coordinate). This means that reads at the start and end of the qseq are from the extreme edges of the tile which have much lower image quality. What you are observing in these qseq files is perfectly normal and expected.

It is also likely that the fastq file they typically send you only contains filter passed reads whereas the qseq files contain all reads. Reads like the one shown would not end up in the filter passing fastq file.

simonandrews · 08-27-2010, 05:08 AM

Incidentally, you can request that your sequencing service provide fastq files still. The latest Illumina pipeline doesn't generate them by default, but there's an option to turn them back on.

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08-26-2010, 09:27 AM	#1
brentp Member Location: salt lake city, UT Join Date: Apr 2010 Posts: 72	qseq files pattern of reads with no sequence i dont believe this is a problem, just something i'm curious about: normally we get our data from a sequencing center that returns a fastq file. this time, we have a bunch of qseq files like: s_3_1_0001_qseq.txt in all of them, the first and last few hundred reads in the file have no sequence (just .'s for the length of the read). as one moves down from the top of the file (or up from the bottom), it starts that sequence appears on either end of the read, so it will look like: Code: G.TCTG..............................................................ACCCGGA. (and the quality is all B's). going to the center of the file, the reads are all sequence. i'm curious as to why this occurs at the ends of the file. is there some correspondence between position in the file and location on the cell. thanks.

08-26-2010, 09:44 AM	#2
kmcarr Senior Member Location: USA, Midwest Join Date: May 2008 Posts: 1,178	Your guess is correct. Reads are sorted by their x-coordinate (but apparently there's no sorting by y-coordinate). This means that reads at the start and end of the qseq are from the extreme edges of the tile which have much lower image quality. What you are observing in these qseq files is perfectly normal and expected. It is also likely that the fastq file they typically send you only contains filter passed reads whereas the qseq files contain all reads. Reads like the one shown would not end up in the filter passing fastq file. Last edited by kmcarr; 08-26-2010 at 09:47 AM.

08-27-2010, 05:08 AM	#3
simonandrews Simon Andrews Location: Babraham Inst, Cambridge, UK Join Date: May 2009 Posts: 871	Incidentally, you can request that your sequencing service provide fastq files still. The latest Illumina pipeline doesn't generate them by default, but there's an option to turn them back on.