Hi all!
Happy 2013!
I am having a bit of an issue with generating consensus sequences from an Alignment BAM file.
The alignment was done with Stampy. It is a mix of single end and paired end data. Looking at the BAM in IGV shows me I have good coverage across the board and reasonable mapping qualites.
I then do the routine samtools mpileup (using -u -f -r paramters; also tried the -A); then bcftools view (- c -g ); followed by conversion to fastq and ultimately fasta.
In my consensus fasta however; I have an inordinate amount of Ns; even in positions; where I can clearly see correct alignment/mapping.
Does anyone know when mpileup is calling an N and what I might be doing wrong?
Funnily enough; I have aligned the exact same data with BWA and done the same process; even though BWA maps less reads properly; with lower mapping quality and coverage; calling the consensus only spits out Ns if there is either no coverage at a given position or there is already an N in the reference.
So, why is this not working from my stampy-generated BAM?
Argh!
Any other recommendations for tools to generate consensus sequences?
Happy 2013!
I am having a bit of an issue with generating consensus sequences from an Alignment BAM file.
The alignment was done with Stampy. It is a mix of single end and paired end data. Looking at the BAM in IGV shows me I have good coverage across the board and reasonable mapping qualites.
I then do the routine samtools mpileup (using -u -f -r paramters; also tried the -A); then bcftools view (- c -g ); followed by conversion to fastq and ultimately fasta.
In my consensus fasta however; I have an inordinate amount of Ns; even in positions; where I can clearly see correct alignment/mapping.
Does anyone know when mpileup is calling an N and what I might be doing wrong?
Funnily enough; I have aligned the exact same data with BWA and done the same process; even though BWA maps less reads properly; with lower mapping quality and coverage; calling the consensus only spits out Ns if there is either no coverage at a given position or there is already an N in the reference.
So, why is this not working from my stampy-generated BAM?
Argh!
Any other recommendations for tools to generate consensus sequences?
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