<p>If you have reasonable coverage with HiFi (30X per haplotype), then there usually isn't any reason to also use Illumina. My understanding is that Illumina can make the assembly worse if you try to polish with it, because it can erroneously induce haplotype switching errors.</p>
<p>I would use <a href="https://genome.cshlp.org/content/early/2020/08/14/gr.263566.120.full.pdf+html" rel="nofollow noreferrer">HiCanu</a> or <a href="https://www.nature.com/articles/s41592-020-01056-5" rel="nofollow noreferrer">HiFiAsm</a>, depending on whether you expect significant heterozygosity, and also assuming that you have non-trivial HiFi coverage.</p>
<p>If your HiFi coverage is very low (&lt;15X per haplotype, maybe), you might consider going back to subreads, assembling those, and polishing with Pilon and your Illumina reads, or one of the workflows that you have found. But otherwise I don't think the Illumina will be very useful. If you had Hi-C reads you could use them directly in HiFiAsm to assist phasing and scaffolding, but I'm assuming that your Illumina is shotgun.</p>
<p>I know it's a PB promo material, but you see that PB promotes HiFi as <a href="https://www.pacb.com/wp-content/uploads/Trade-Up-to-HiFi-Sequencing-on-the-Sequel-IIe-System_North-America-Sales.pdf" rel="nofollow noreferrer">not needing polishing</a>, as <a href="https://academic.oup.com/gigascience/article/9/12/giaa123/6034784" rel="nofollow noreferrer">HiFi has accuracy &gt;99%, so shotgun Illumina is not going to do much</a>. There is some argument for still polishing with your original long reads.</p>
<p>I can recommend any of the linked papers for ideas or further explanation.</p>

CC BY-SA 4.0
2021-10-24T14:57:31.583Z
Unfortunately, my HiFi coverage isn't that great, (unexpectedly large genome). However, my shotgun short read dataset is large and good quality, hence the question.
bishopia
CC BY-SA 4.0
2021-10-24T18:41:46.997Z
@bishopia Hmmmm I see what you mean. In that case, subread assembly+Illumina polishing might be your best bet. It would still be helpful to know more specifics (e.g. coverage, read length profile of each dataset: CCS, subread, ILMN). I am pretty skeptical of the usefulness of Illumina contigs as the basis of assemblies of large/complex/heterozygous genomes, compared to even low-coverage PB data.
Maximilian Press
CC BY-SA 4.0
2021-10-23T03:45:34.477Z
2021-10-23T03:45:34.477Z
Maximilian Press
<p>I would like to perform <em>de novo</em> genome assembly on a diploid microalgal strain.</p>
<p>I have two datasets:</p>
<ul>
<li>PacBio CCS/HiFi reads, low coverage.</li>
<li>Illumina PE 2x150 (standard shotgun)</li>
</ul>
<p>Does anybody have any software recommendations for how best to use both in a hybrid <em>de novo</em> assembly?</p>
<p>Most options out there are set up for Pacbio CLR reads, not Pacbio CCS/HiFi, for using in hybrid assembly. Perhaps I should assemble short read contigs first then combine with <a href="https://github.com/adigenova/wengan#pacccs-experimental" rel="nofollow noreferrer">wengan</a>, or go back to the PacBio subreads instead of CCS to use in a hybrid method?</p>

CC BY-SA 4.0
2021-10-23T14:39:46.390Z
Please clarify your specific problem or provide additional details to highlight exactly what you need. As it's currently written, it's hard to tell exactly what you're asking.
Community
<assembly><software-recommendation><illumina><pacbio>
How to de novo hybrid assemble with Pacbio CCS and Illumina PE reads
# How to de novo hybrid assemble with Pacbio CCS and Illumina PE reads

(Asked by: username_0 on 2021-10-22)

I would like to perform *de novo* genome assembly on a diploid microalgal strain.

I have two datasets:

* PacBio CCS/HiFi reads, low coverage.
* Illumina PE 2x150 (standard shotgun)

Does anybody have any software recommendations for how best to use both in a hybrid *de novo* assembly?

Most options out there are set up for Pacbio CLR reads, not Pacbio CCS/HiFi, for using in hybrid assembly. Perhaps I should assemble short read contigs first then combine with [wengan](https://github.com/adigenova/wengan#pacccs-experimental), or go back to the PacBio subreads instead of CCS to use in a hybrid method?

**Tags:** assembly, software-recommendation, illumina, pacbio
> **Comments:**
> **username_1**: Please clarify your specific problem or provide additional details to highlight exactly what you need. As it's currently written, it's hard to tell exactly what you're asking.  

---

## 1 answers

### Answer by username_2 on 2021-10-23. Score: 1

If you have reasonable coverage with HiFi (30X per haplotype), then there usually isn't any reason to also use Illumina. My understanding is that Illumina can make the assembly worse if you try to polish with it, because it can erroneously induce haplotype switching errors.

I would use [HiCanu](https://genome.cshlp.org/content/early/2020/08/14/gr.263566.120.full.pdf+html) or [HiFiAsm](https://www.nature.com/articles/s41592-020-01056-5), depending on whether you expect significant heterozygosity, and also assuming that you have non-trivial HiFi coverage.

If your HiFi coverage is very low (<15X per haplotype, maybe), you might consider going back to subreads, assembling those, and polishing with Pilon and your Illumina reads, or one of the workflows that you have found. But otherwise I don't think the Illumina will be very useful. If you had Hi-C reads you could use them directly in HiFiAsm to assist phasing and scaffolding, but I'm assuming that your Illumina is shotgun.

I know it's a PB promo material, but you see that PB promotes HiFi as [not needing polishing](https://www.pacb.com/wp-content/uploads/Trade-Up-to-HiFi-Sequencing-on-the-Sequel-IIe-System_North-America-Sales.pdf), as [HiFi has accuracy >99%, so shotgun Illumina is not going to do much](https://academic.oup.com/gigascience/article/9/12/giaa123/6034784). There is some argument for still polishing with your original long reads.

I can recommend any of the linked papers for ideas or further explanation.

> **Comments:**
> **username_0**: Unfortunately, my HiFi coverage isn't that great, (unexpectedly large genome). However, my shotgun short read dataset is large and good quality, hence the question.  
> **username_2**: @bishopia Hmmmm I see what you mean. In that case, subread assembly+Illumina polishing might be your best bet. It would still be helpful to know more specifics (e.g. coverage, read length profile of each dataset: CCS, subread, ILMN). I am pretty skeptical of the usefulness of Illumina contigs as the basis of assemblies of large/complex/heterozygous genomes, compared to even low-coverage PB data.  

---

