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Old 03-02-2018, 12:45 AM   #1
meriem
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Location: morocco

Join Date: May 2016
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Question can I remove the N's in my sequence?

Hello,

I'm having a problem with mine assembly, I have a bacterial genome of 3.7bp assembled into 24 contigs, which was part of an under submission research, the reviewer gave me as a feedback that I should have a gap free sequence; Unfrontutly I can't not re-do the sequencing or use Pacbio. so I used ragout for reference-assisted assembly to improve the quality of my assembly; I got 1 scaffold of 3.7 bp, but a lot of ns (40000 n), how can I fix this problem? is this software reliable since I got some small fragment left? can I remove the ns without affecting the sequence?

thank you
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Old 03-08-2018, 09:56 AM   #2
cstack
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Just so we are using the same terminology: An assembly gap is typically represented by a series of N's connecting two contigs; the product of 2+ contigs connected by gaps is referred to as a scaffold.

Your 24 original contigs contained no gaps (Ns), correct? I would confirm this first

I have never used Ragout, but it seems to order and connect contigs based on a reference genome; in the process of connecting contigs, gap characters (Ns) are added. There is nothing atypical about this -- Without looking over your manuscript, it is difficult to say why the review took issue with the gaps.

Anyhow, you can use
Code:
seqtk cutN -n 1 ragout.fasta
to break up the single, 3.7(M?)bp scaffold produced by ragout into a set of gap-free contigs.
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Old 03-09-2018, 03:39 AM   #3
meriem
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Quote:
Originally Posted by cstack View Post
Just so we are using the same terminology: An assembly gap is typically represented by a series of N's connecting two contigs; the product of 2+ contigs connected by gaps is referred to as a scaffold.

Your 24 original contigs contained no gaps (Ns), correct? I would confirm this first

I have never used Ragout, but it seems to order and connect contigs based on a reference genome; in the process of connecting contigs, gap characters (Ns) are added. There is nothing atypical about this -- Without looking over your manuscript, it is difficult to say why the review took issue with the gaps.

Anyhow, you can use
Code:
seqtk cutN -n 1 ragout.fasta
to break up the single, 3.7(M?)bp scaffold produced by ragout into a set of gap-free contigs.
Thank you for your reply,
My 24 contigs doesn't contain any N's.
Im' doing a comparative genomic analysis of 2 strains from an extrem envirenement, I got this msg from the reviewer " In that the strains on which this Genome Report is based can be grown in culture, it is necessary to produce gap-free sequences for both strains, since the data would be more generally useful for the community at large. Given the high coverage that is already available, it would probably be necessary to take a more direct approach (rather than increasing coverage)."

If I Use this commande I will go back to my initial contigs, No?
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