<p>The number of contigs and total assembly size you have suggest that there was probably more in the sequencing run than a single virus strain. Does the total assembly length correspond to expected genome size of the reference strain? Have you tried just to blast some your contigs? Could you have a contamination or something?</p>

<p>Maybe you can just sort out the contigs by the reference using something like <a href="https://www.sanger.ac.uk/science/tools/artemis" rel="nofollow noreferrer">artemis</a> or <a href="http://bioinf.spbau.ru/quast" rel="nofollow noreferrer">quast</a> compares your assembly to reference and showing nice stats (just to be sure that your assembly makes sense so far). Or you could use directly <a href="https://github.com/mummer4/mummer" rel="nofollow noreferrer">MUMmer</a> (which the alignment tool that is running on the backend of of Medusa and Quast).</p>

<p>This is not directly answering your question "How to scaffold", however I think you should first figure out if your contigs make sense before you scaffold them. Also sorting your contigs by artemis using reference is practically the same thing as scaffolding since you won't know the sequence in between in either of the cases.</p>

CC BY-SA 4.0
2018-07-23T21:28:31.117Z
You are right. There is a lot of host genome contamination. I blasted all the contigs against reference genome and only about 10 contigs had significant match with the reference. I was looking for some tool that could take these 10 contigs, re-order them and join them together or add NNNNs if the sequence is missing between the contigs.
LoveVirology
CC BY-SA 4.0
2018-07-23T21:42:08.387Z
And do you know the host genome? If yes, you could map everything to both host and the reference and then kick out everything that maps to host and scaffold just non-host contigs.
Kamil S Jaron
CC BY-SA 4.0
2018-07-23T23:44:43.350Z
I know the host genome because it is cell-cultured virus. And I am actually looking for some tools that could actually do what you have mentioned.
LoveVirology
CC BY-SA 4.0
2018-07-24T06:25:36.727Z
I would personally just run MUMmer of the assembly vs catinated reference of the host and related strain. MUMmer allows secondary alignments as well so you will be able to find also regions that are similar between the both, then parse the output in python/R/whateveryoulike to create a list of scaffolds you want keep/remove and use it to filter out our reference (a script to do that could look like this https://github.com/KamilSJaron/generic_genomics/blob/master/fasta2extract_by_list_of_headers.py). I should probably add this to the answer.
Kamil S Jaron
CC BY-SA 4.0
2018-07-26T02:19:45.363Z
Thanks Kamil. I will try that script. I also found that Viral NGS Pipeline from broad institute can actually do this. There is a python script called order_and_orient which serves this purpose. Here is the link https://viral-ngs.readthedocs.io/en/latest/assembly.html
LoveVirology
CC BY-SA 4.0
2018-07-23T19:08:13.030Z
2018-07-23T19:08:13.030Z
Kamil S Jaron
<p>I found that Viral NGS Pipeline from broad institute can actually do this. There is a python script called <strong>order_and_orient</strong> which serves this purpose. Here is the link  <a href="https://viral-ngs.readthedocs.io/en/latest/assembly.html" rel="nofollow noreferrer">viral ngs assembly</a></p>

CC BY-SA 4.0
2018-07-26T02:23:08.203Z
2018-07-26T02:23:08.203Z
LoveVirology
<p>What you mean is assembly based scaffolding, as opposed to using reads with long distance information such as mate pair/long jumping distance to scaffold.</p>

<p>You could reduce your contig file to just the longest contigs, since they have the most information. Probably 156MB of data is overkill for your virus. </p>

<p>There are other programs out there. </p>

<p><a href="https://github.com/ksahlin/BESST" rel="nofollow noreferrer">https://github.com/ksahlin/BESST</a></p>

<p><a href="https://github.com/institut-de-genomique/MaGuS" rel="nofollow noreferrer">https://github.com/institut-de-genomique/MaGuS</a></p>

<p>Your mileage will vary.</p>

CC BY-SA 4.0
2018-07-20T17:55:20.663Z
I tried that. I took five largest contigs. The output is not good. I will try the tools that you provided. Thank you.
LoveVirology
CC BY-SA 4.0
2018-08-14T13:22:28.497Z
Isn't BESST only for read-based scaffolding?
Kamil S Jaron
CC BY-SA 4.0
2018-07-19T11:38:55.660Z
2018-07-19T11:38:55.660Z
Colin Davenport
<p>I have assembled a virus genome using <a href="https://github.com/sebhtml/ray" rel="nofollow noreferrer">Ray</a> resulting in approximately 5000 contigs. Now I want to build a scaffold of those contigs to get the full genome (I am aware of the fact that there might be some gaps in the genome).</p>
<p>I  found one tool called <a href="https://github.com/combogenomics/medusa" rel="nofollow noreferrer">Medusa</a> to do this. Unfortunately, this tool cannot take a file above 50 MB and my contig file is about 156 MB.</p>
<p>Are there any other tools that can perform reference based scaffolding?</p>
<p>It would be great if anyone knows about genome scaffolding packages in Bioconda.</p>

CC BY-SA 4.0
2018-07-19T12:02:16.957Z
Any genomics tool that doesn't work over 50MB data is meaningless. Please delete Medusa.
SmallChess
CC BY-SA 4.0
2018-07-23T18:52:49.673Z
@SmallChess if it's designed for viral genomes, why not.
Kamil S Jaron
<ngs><covid-19><sars-cov-2><reference-genome><scaffold>
Genome scaffolding
# Genome scaffolding

(Asked by: username_0 on 2018-07-17)

I have assembled a virus genome using [Ray](https://github.com/sebhtml/ray) resulting in approximately 5000 contigs. Now I want to build a scaffold of those contigs to get the full genome (I am aware of the fact that there might be some gaps in the genome).

I found one tool called [Medusa](https://github.com/combogenomics/medusa) to do this. Unfortunately, this tool cannot take a file above 50 MB and my contig file is about 156 MB.

Are there any other tools that can perform reference based scaffolding?

It would be great if anyone knows about genome scaffolding packages in Bioconda.

**Tags:** ngs, covid-19, sars-cov-2, reference-genome, scaffold
> **Comments:**
> **username_1**: Any genomics tool that doesn't work over 50MB data is meaningless. Please delete Medusa.  
> **username_2**: @SmallChess if it's designed for viral genomes, why not.  

---

## 3 answers

### Answer by username_2 on 2018-07-23. Score: 2

The number of contigs and total assembly size you have suggest that there was probably more in the sequencing run than a single virus strain. Does the total assembly length correspond to expected genome size of the reference strain? Have you tried just to blast some your contigs? Could you have a contamination or something?

Maybe you can just sort out the contigs by the reference using something like [artemis](https://www.sanger.ac.uk/science/tools/artemis) or [quast](http://bioinf.spbau.ru/quast) compares your assembly to reference and showing nice stats (just to be sure that your assembly makes sense so far). Or you could use directly [MUMmer](https://github.com/mummer4/mummer) (which the alignment tool that is running on the backend of of Medusa and Quast).

This is not directly answering your question "How to scaffold", however I think you should first figure out if your contigs make sense before you scaffold them. Also sorting your contigs by artemis using reference is practically the same thing as scaffolding since you won't know the sequence in between in either of the cases.

> **Comments:**
> **username_0**: You are right. There is a lot of host genome contamination. I blasted all the contigs against reference genome and only about 10 contigs had significant match with the reference. I was looking for some tool that could take these 10 contigs, re-order them and join them together or add NNNNs if the sequence is missing between the contigs.  
> **username_2**: And do you know the host genome? If yes, you could map everything to both host and the reference and then kick out everything that maps to host and scaffold just non-host contigs.  
> **username_0**: I know the host genome because it is cell-cultured virus. And I am actually looking for some tools that could actually do what you have mentioned.  
> **username_2**: I would personally just run MUMmer of the assembly vs catinated reference of the host and related strain. MUMmer allows secondary alignments as well so you will be able to find also regions that are similar between the both, then parse the output in python/R/whateveryoulike to create a list of scaffolds you want keep/remove and use it to filter out our reference (a script to do that could look like this https://github.com/KamilSJaron/generic_genomics/blob/master/fasta2extract_by_list_of_headers.py). I should probably add this to the answer.  
> **username_0**: Thanks Kamil. I will try that script. I also found that Viral NGS Pipeline from broad institute can actually do this. There is a python script called order_and_orient which serves this purpose. Here is the link https://viral-ngs.readthedocs.io/en/latest/assembly.html  

---

### Answer by username_0 on 2018-07-26. Score: 1

I found that Viral NGS Pipeline from broad institute can actually do this. There is a python script called **order\_and\_orient** which serves this purpose. Here is the link [viral ngs assembly](https://viral-ngs.readthedocs.io/en/latest/assembly.html)


---

### Answer by username_3 on 2018-07-19. Score: 1

What you mean is assembly based scaffolding, as opposed to using reads with long distance information such as mate pair/long jumping distance to scaffold.

You could reduce your contig file to just the longest contigs, since they have the most information. Probably 156MB of data is overkill for your virus.

There are other programs out there.

<https://github.com/ksahlin/BESST>

<https://github.com/institut-de-genomique/MaGuS>

Your mileage will vary.

> **Comments:**
> **username_0**: I tried that. I took five largest contigs. The output is not good. I will try the tools that you provided. Thank you.  
> **username_2**: Isn't BESST only for read-based scaffolding?  

---

