Is Dorado, nanopolish, and xpore compatible with each other?

Question

[ward.alafef]

Hi this is my first post here, budding bioinformatician.

I have basecalled pod5 files and aligned them at the same time using Dorado while running one of its modification detection models. Using my output BAMs I ran samtools :

samtools view -b -F 0x904 --threads 40 "$input_bam" > "${prefix}_primary.bam"

samtools view -b -q 7 -@ 40 "${prefix}_primary.bam" > "${prefix}_primary_mapq7.bam"

samtools sort --threads 40 -o "${prefix}_primary_mapq7_sorted.bam" "${prefix}_primary_mapq7.bam"

samtools index -@ 40 "${prefix}_primary_mapq7_sorted.bam" 

Can I run nanopolish on the final bam? My aim is to use xpore to do Diferrential RNA modification to analyze change in modification levels b/t wt and KO.

But to run xPore I need to resquiggle with nanopolish and some sources are saying that it only works with guppy which is a precursor of dorado.

Answer 1

[GenoMax]

Use a current program like modkit (LINK) provided by ONT.

As noted on the nanopolish site https://github.com/jts/nanopolish?tab=readme-ov-file#a-note-on-r10-support the program does not support R10.4 flowcells (which is likely what you have, if your data is recent).

dorado is actively developed recent basecaller that is excellent (especially in super accuracy mode). If you have not done this you can try that. You can also do read error correction with dorado correct (which uses SUP reads). Looks like there is also dorado polish.