CLC and SNP discovery

extari · 04-11-2011, 06:27 AM

Hi there,

I am using CLC 4.5 software for doing a de novo assembly and SNP discovery.
I have already perform my de novo assembly, and I have mapped my reads to contigs with the main objective of SNP discovery.
This software (CLC) has the option "SNP Detection", I have done it and I have discovered SNPs (after filtering).
Does anyone of all know how can I get my consensus sequences with the ambiguities in one fasta file, or txt format?

I mean, imagine this sequence with this SNP (*), and these ambiguities (-)
5` ... A G G G A T A G C C C G A T A G G G T A C A G G T T A ... 3`
- * - -
When I try to save my consensus sequence as a txt file, I can only save as one of these:
a) if in the assembly selected VOTE (for ambiguities):
5` ... A G G G A T A G C C C G A T A G G G T A C A G G T T A ... 3`

b) if in the assembly selected Unknown (for ambiguities):
5` ... A G G G N T A G C C N G A T N G G G T A N A G G T T A ... 3`

c) if in the assembly selected Ambiguity (for ambiguities):
5` ... A G G G A R A G C C C R A T M G G G T A Y A G G T T A ... 3`

But the one I want to save is this one: 5` ... A G G G A T A G C C C R A T A G G G T A C A G G T T A ... 3` (with the only annotated SNP).

Could you help me, please???

Thank you!

flipwell · 04-11-2011, 04:46 PM

I haven't tried it, but during SNP detection you can select to annotate the consensus with SNPs or not - if you choose vote for ambiguities during alignment then annotate with SNPs during detection I'm guessing it should give you the result you are after

extari · 04-13-2011, 06:45 AM

Hi ya,

I have tried to do what you have told me, but that one is not the solution, look what happens when I do that (see the attachment).
The SNP is annotated, but I cannt have the fasta file as ambiguities (R) or as [A/G], what is the thing I would like, there must be a way to do that, but, how?

NextGenSeq · 04-13-2011, 08:55 AM

That's a terrible alignment. You should increase the gap penalty.

extari · 04-13-2011, 11:01 AM

no, don't worry about the aligment, that was a bad example.
the question is how can i have the fasta file with the polymorphic site?

szilva · 04-13-2011, 11:20 PM

If you point to the sequence name on the left side of the panel and right click on the name, you should get a menu like "Open Copy of Sequence" . Once you have got a copy, you can export that into FASTA.

extari · 04-14-2011, 01:24 AM

Yes, that is true, but the problem persists, in this way I cannot have the polymorphism annotation in the sequence (as "R" or "[A/G]") in the FASTA file.
I don't know how to do this, I am desperate!!!

sklages · 04-14-2011, 02:10 AM

what does CLC recommend? AFAIK their support is quite helpful.

extari · 04-14-2011, 02:13 AM

In the user guide there is nothing about this issue.
I have sent them an e-mail, I am waiting for response. If I can solve this issue, I will tell you, but I think there is no way to do this...

extari · 04-15-2011, 02:32 AM

Hello all,

CLC support has answered to my question in a e-mail. They have told me this: "I'm afraid there is no way to export fasta sequence with ambiguities or IUPAC letters from the CLC software. However, if you have chosen to annotate your Consensus, and/or your Reference sequence, you could then export to GFF or Genbank format. The SNP variants that have been called will appear in the annotations. At that point though, you would have to script something to generate files of the format that suits you needs."

So, there is no way to do this...

If anyone knows about these type of script, please, let me know.

Thank you!

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04-11-2011, 06:27 AM	#1
extari Junior Member Location: Bilbao (Spain) Join Date: Mar 2011 Posts: 9	CLC and SNP discovery Hi there, I am using CLC 4.5 software for doing a de novo assembly and SNP discovery. I have already perform my de novo assembly, and I have mapped my reads to contigs with the main objective of SNP discovery. This software (CLC) has the option "SNP Detection", I have done it and I have discovered SNPs (after filtering). Does anyone of all know how can I get my consensus sequences with the ambiguities in one fasta file, or txt format? I mean, imagine this sequence with this SNP (), and these ambiguities (-) 5` ... A G G G A T A G C C C G A T A G G G T A C A G G T T A ... 3` - - - When I try to save my consensus sequence as a txt file, I can only save as one of these: a) if in the assembly selected VOTE (for ambiguities): 5` ... A G G G A T A G C C C G A T A G G G T A C A G G T T A ... 3` b) if in the assembly selected Unknown (for ambiguities): 5` ... A G G G N T A G C C N G A T N G G G T A N A G G T T A ... 3` c) if in the assembly selected Ambiguity (for ambiguities): 5` ... A G G G A R A G C C C R A T M G G G T A Y A G G T T A ... 3` But the one I want to save is this one: 5` ... A G G G A T A G C C C R A T A G G G T A C A G G T T A ... 3` (with the only annotated SNP). Could you help me, please??? Thank you!

04-15-2011, 02:32 AM	#10
extari Junior Member Location: Bilbao (Spain) Join Date: Mar 2011 Posts: 9	CLC support has answered Hello all, CLC support has answered to my question in a e-mail. They have told me this: "I'm afraid there is no way to export fasta sequence with ambiguities or IUPAC letters from the CLC software. However, if you have chosen to annotate your Consensus, and/or your Reference sequence, you could then export to GFF or Genbank format. The SNP variants that have been called will appear in the annotations. At that point though, you would have to script something to generate files of the format that suits you needs." So, there is no way to do this... If anyone knows about these type of script, please, let me know. Thank you!

04-11-2011, 04:46 PM	#2
flipwell Member Location: Queensland Join Date: Feb 2011 Posts: 14	I haven't tried it, but during SNP detection you can select to annotate the consensus with SNPs or not - if you choose vote for ambiguities during alignment then annotate with SNPs during detection I'm guessing it should give you the result you are after

04-13-2011, 08:55 AM	#4
NextGenSeq Senior Member Location: USA Join Date: Apr 2009 Posts: 482	That's a terrible alignment. You should increase the gap penalty.

04-13-2011, 11:01 AM	#5
extari Junior Member Location: Bilbao (Spain) Join Date: Mar 2011 Posts: 9	no, don't worry about the aligment, that was a bad example. the question is how can i have the fasta file with the polymorphic site?

04-13-2011, 11:20 PM	#6
szilva Member Location: Hungary Join Date: Aug 2009 Posts: 16	If you point to the sequence name on the left side of the panel and right click on the name, you should get a menu like "Open Copy of Sequence" . Once you have got a copy, you can export that into FASTA.

04-14-2011, 01:24 AM	#7
extari Junior Member Location: Bilbao (Spain) Join Date: Mar 2011 Posts: 9	Yes, that is true, but the problem persists, in this way I cannot have the polymorphism annotation in the sequence (as "R" or "[A/G]") in the FASTA file. I don't know how to do this, I am desperate!!!

04-14-2011, 02:10 AM	#8
sklages Senior Member Location: Berlin, DE Join Date: May 2008 Posts: 628	what does CLC recommend? AFAIK their support is quite helpful.

04-14-2011, 02:13 AM	#9
extari Junior Member Location: Bilbao (Spain) Join Date: Mar 2011 Posts: 9	In the user guide there is nothing about this issue. I have sent them an e-mail, I am waiting for response. If I can solve this issue, I will tell you, but I think there is no way to do this...