I have sequenced a bacterial genome with a GridIon from ONT. Basically what I want to check is whether or not trimming 50 bps at the beginning of the reads will improve alignment against the reference genome and ultimately the call of a consensus sequence.
For that what I have done so far is:
- concatenated all the fastqs generated from the Gridion into a single (very large) fastq file:
- aligned this concatenated fastq against the reference genome
~/Downloads/minimap2-2.17_x64-linux/minimap2 -ax map-ont ref_genbank/ref_genbank.fasta ref494_cat_all.fastq > ref494_aligned_trim50.sam
- Performed the general
samtoolssteps to covert sam to bam, index and sort
Then, in order to get the consensus sequence I first try to generate a
.vcf.gz file, according to this tutorial
bcftools mpileup -f ref_genbank/ref_genbank.fasta ref494_aligned.sorted.bam | bcftools call -mv -Oz -o calls.vcf.gz
Immediatelly after calling the command above I got:
Note: none of --samples-file, --ploidy or --ploidy-file given, assuming all sites are diploid [mpileup] 1 samples in 1 input files
Then, after a minute or two, I got:
Failed to open -: unknown file type
I'm pretty sure that there's no issue with
.bam file, as I could visualise the alignment with IGV or tablet.
Does anyone know what the issue can be?