why consed finds discrepancies with masked n sequence

alig · 03-05-2009, 05:55 PM

Hi,

I have just received our first Illumina data & I have assembled it into Consed. Then I searched for high quality discrepancies as am looking for variants.

But also in the list brought up it has discrepancies with 'n' (masked) sequence in the RefSeq.

Why would the program compare the sequences to regions of N's . I would have thought it would only be looking for differences between the 4 bases.

How can I screen these out to reduce my long list of discrepancies?

Thanks alig

basickler · 03-12-2009, 09:29 AM

If you're using Phrap/Consed you might want to think about using something like MAQ or similar programs as Phrep/Consed will not do well with short read sequencing (doesn't really do well on anything but sanger BAC re-sequencing/assembly without tweaking it a lot really). If you're working on de-novo stuff and don't have a reference try velvet or euler-sr.
Cheers,

Brad

sklages · 03-12-2009, 10:54 AM

Consed uses cross_match to align the reads (454,illumina) against a reference sequence,
phrap is not invoked. If you are "working" with the alignment, e.g. some kind of finishing, then
you are somewhat dependent on programs like consed.

But I agree that consed / cross_match is not doing that well on that, mainly because of the
problem mentioned and as well as it (still) lacks some proper consensus recalculation after
(short) reads have been aligned.

So if you're just looking for discrepancies, I'd also recommend programs like MAQ or
Mosaik Assembler (from the Marth Lab) which is well suited for SNP searches as well.

hth,
Sven

bioinfosm · 06-12-2009, 12:32 PM

cross match seems pretty sensitive to aligning short reads correctly. It is a matter of obtaining SNP calls from it and then compare.

The discrepancies reported by consed need to go through some sort of filtering to be able to get good quality snp calls.. anyone worked with this?

perhaps cross match mapped reads -> maq map format -> maq SNP calling?

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03-05-2009, 05:55 PM	#1
alig Member Location: adelaide Join Date: Sep 2008 Posts: 43	why consed finds discrepancies with masked n sequence Hi, I have just received our first Illumina data & I have assembled it into Consed. Then I searched for high quality discrepancies as am looking for variants. But also in the list brought up it has discrepancies with 'n' (masked) sequence in the RefSeq. Why would the program compare the sequences to regions of N's . I would have thought it would only be looking for differences between the 4 bases. How can I screen these out to reduce my long list of discrepancies? Thanks alig

03-12-2009, 09:29 AM	#2
basickler Member Location: San Diego Join Date: Mar 2009 Posts: 26	If you're using Phrap/Consed you might want to think about using something like MAQ or similar programs as Phrep/Consed will not do well with short read sequencing (doesn't really do well on anything but sanger BAC re-sequencing/assembly without tweaking it a lot really). If you're working on de-novo stuff and don't have a reference try velvet or euler-sr. Cheers, Brad

03-12-2009, 10:54 AM	#3
sklages Senior Member Location: Berlin, DE Join Date: May 2008 Posts: 628	Consed uses cross_match to align the reads (454,illumina) against a reference sequence, phrap is not invoked. If you are "working" with the alignment, e.g. some kind of finishing, then you are somewhat dependent on programs like consed. But I agree that consed / cross_match is not doing that well on that, mainly because of the problem mentioned and as well as it (still) lacks some proper consensus recalculation after (short) reads have been aligned. So if you're just looking for discrepancies, I'd also recommend programs like MAQ or Mosaik Assembler (from the Marth Lab) which is well suited for SNP searches as well. hth, Sven

06-12-2009, 12:32 PM	#4
bioinfosm Senior Member Location: USA Join Date: Jan 2008 Posts: 482	cross match seems pretty sensitive to aligning short reads correctly. It is a matter of obtaining SNP calls from it and then compare. The discrepancies reported by consed need to go through some sort of filtering to be able to get good quality snp calls.. anyone worked with this? perhaps cross match mapped reads -> maq map format -> maq SNP calling?