Flye Assembler Nano-Raw vs Nano-HQ options

Question

[Umer]

Hello reader.

Asking this question again because i couldn’t find a proper answer or suggestion.

I have some nanopore sequenced fungal sample. Sequenced with R10.4 cell and LSK114 kit.

Basecalling was performed using dorado and from fastQ files i found basecall_model_version_id=dna_r10.4.1_e8.2_400bps_hac@v4.2.0

Median Q-score across all samples is between 16-17

Median Read-length across all samples is between 7000-9000 in all samples less than 3% reads are actually >Q20.

I previously used Flye genome assembler version: 2.9.5 with nano-raw option for assembly generation but now i see some papers which have data sequenced with same nanopore cell and chemistry and they used nano-hq option.

This makes me confused on which option should be used for my data.

Can you share some insights on this. Will be a great assistance.

Previous command I used

flye --nano-raw NP01_chopper.fastq --out-dir ./flye_assemblies/NP01/ --threads 45 --genome-size 60m --iterations 2 --scaffold

Thank you.

Answer 1

[shelkmike]

First, I recommend to do basecalling in sup mode instead of hac. You'll likely have a better assembly with sup reads.
Regarding your question: I suggest to try both options and see which one will result in a better assembly. I suppose that the assemblies will be of similar quality, but the assembly with --nano-hq will take less time.
Also, in addition to Flye it's probably worth trying the latest release of Hifiasm with option "--ont".

[Umer]

Hello, Thanks for the response.

which dorado version to use as there are two different latest builds. v0.7.4 and v0.8.3

also, will Hifiasm work for haploid fungal genomes ? as far as i know it is built for diploid genomes. I have tested canu and nextDenovo but flye yeilded better results.

[shelkmike]

Hifiasm can assemble haploid genomes. Use "--n-hap 1".

[GenoMax]

Use the latest. You will need access to the original POD5/fast5 files.

BTW Is this a new dataset? You have been asking questions about fungal assemblies for a while.

[Umer]

Its the same fungal dataset. but i get confused when i go through different papers. about the analysis steps. now i saw this Nano-HQ option in one paper for same species and got me confused if i am doing it all wrong.