[validation] How to arrange the mtDNA after assembly with mitoHiFi from hifiasm

[Isoris]

Hello,

I have assembled a mitogenome with mitoHiFi using a reference genome from a closely related species, starting from Hifiasm contigs, I could get the final.mitogenome.fasta , then I remapped HiFi reads as follows:

``
minimap2 -t 32 -ax map-hifi --secondary=no final_mitogenome.fasta ../../../00-raw_data/hifi/CM_M_HIFI_None_reads.fastq.gz | samtools view -@ 32 -bhS - | samtools sort -@ 32 -o final_mitogenome.fasta.hifi.aligned.sorted.bam

index the bam files with samtools index

for file in *.bam
do
samtools index -@ 16 --bai ${file} -o ${file}.bai
bedtools genomecov -ibam ${file} -bg > ${file}.coverage.bedgraph.bed
done

igv -g final_mitogenome.fasta final_mitogenome.fasta.hifi.aligned.filtered.sorted.bam
``
however, I could see that there are some inconsistencies in the sequence:

1. for instance there are breakpoints with no evidence of support from read coverage (see below):

2. And also regions which seems to be heterozygous:

3. what are those repeats?

Do you have any idea why is it happening?

Below is the gb annotation file:

final_mitogenome.zip

in the region from 700- 8000 there is no annotations for any genes or anything. I can see tRNAs from 0-700 and from 11,000 to the end ~26,000

Thank you in advance for your answers and help :)

Quentin

[Isoris]

Here are the annotation results:

1. Contigs
   
2. Final mtDNA
   

Could suggest some next steps? also do you think I rather use the ONT hifiasm asm as input or start from the ONT reads instead? ?

Thank you for your help :)

[Isoris]

Do you think that there could be contamination in the nanopore data?