Human leukocyte antigen (HLA) genes are critical to humans' adaptive immune response. Because of this, correctly matching the genotypes of these genes between donor and recipient is critical for success in organ transplantation. The most routinely typed HLA genes are HLA-A, HLA-B, HLA-C and HLA-DRB1. HLA sequencing technologies have traditionally focused on the most polymorphic regions encoding the peptide-binding groove that binds to HLA antigens, i.e., exons 2 and 3 for class I genes and exon 2 for class II genes. The antigen-binding groove region of HLA molecules is the focus point of the T-cell receptor and mediates transplant rejection and graft-versus-host diseases (GVHD). For HLA-A, HLA-B and HLA-C, exons 2 and 3 are the most polymorphic exons in the human genome, and are the focus of HLA typing products.
Current DNA-based technologies independently type exons 2 and 3 for HLA-A, HLA-B and HLA-C. This is usually done by DNA hybridization. To gain direct information on the formed proteins, testing can also be done serologically. An alternative method to these is typing by sequencing (sequencing based typing, or SBT). SBT has the potential to provide higher information content than serological or hybridization methods that can, in turn, provide prognostic and diagnostic advantages by avoiding typing ambiguities that plague other methods. However, existing sequencing technologies typically do not have read lengths that allow for complete sequencing of both exons together and/or require cloning each haplotype separately prior to sequencing to resolve within-exonic phase. Also, high error rates are associated with many sequencing technologies. As such, there remains a great need for improved HLA haplotyping methodologies.