I have HLA typed whole genome sequencing data using [HLA-VBSeq][1]. Reads were first aligned to hg19 with BWA-MEM, then reads aligning to all HLA loci and unmapped reads were extracted from the bam file using samtools. The extracted reads were then combined and re-aligned to the collection of all the genomic HLA allele sequences in the IMGT/HLA database using BWA-MEM, allowing for multiple alignments to the reference sequences.

Now, I would like to subset the reads of a specific HLA-E gene allele, however, using `samtools view input.bam HLA:HLA00936`, I see that all of the sequences have a MAPQ score of 0. I know that this is specific to BWA and indicates that the reads are mapping to multiple locations. Does this mean that the alignment results for this allele are insignificant and should be disregarded? The researcher interested in this allele would like to know the HLA-E sequences for gene editing purposes, but I feel as though the coverage is far too low. Any input or advice is much appreciated.





  [1]: http://nagasakilab.csml.org/hla/
If you have multiple inferred HLA-E alleles, then it is expected to have most reads mapping to multiple sequences, because HLA-E is one of the least polymorphic HLA genes. Many sequence segments will be identical between the alleles. You can get the [nucleotide sequences of the HLA-E alleles][1] as a FASTA file and see how similar they are with the multiple sequence alignment tool [MUSCLE][2] (choose HTML output format). You must have some reads mapping uniquely to the allele you are investigating because you inferred that the allele exists from the data.

  [1]: ftp://ftp.ebi.ac.uk/pub/databases/ipd/imgt/hla/fasta/E_nuc.fasta
  [2]: http://www.ebi.ac.uk/Tools/msa/muscle/