title: An improved and validated RNA HLA class I SBT approach for obtaining full length coding sequences.
abstract: The functional relevance of human leukocyte antigen (HLA) class I allele polymorphism beyond exons 2 and 3 is difficult to address because more than 70% of the HLA class I alleles are defined by exons 2 and 3 sequences only. For routine application on clinical samples we improved and validated the HLA sequence-based typing (SBT) approach based on RNA templates, using either a single locus-specific or two overlapping group-specific polymerase chain reaction (PCR) amplifications, with three forward and three reverse sequencing reactions for full length sequencing. Locus-specific HLA typing with RNA SBT of a reference panel, representing the major antigen groups, showed identical results compared to DNA SBT typing. Alleles encountered with unknown exons in the IMGT/HLA database and three samples, two with Null and one with a Low expressed allele, have been addressed by the group-specific RNA SBT approach to obtain full length coding sequences. This RNA SBT approach has proven its value in our routine full length definition of alleles.
authors.affiliation: Department of Transplantation Immunology, Tissue Typing Laboratory, Maastricht University Medical Centre, Maastricht, The Netherlands.
authors.forename: K E H
authors.initials: KE
authors.lastname: Gerritsen
authors.forename: T I
authors.initials: TI
authors.lastname: Olieslagers
authors.forename: M
authors.initials: M
authors.lastname: Groeneweg
authors.forename: C E M
authors.initials: CE
authors.lastname: Voorter
authors.forename: M G J
authors.initials: MG
authors.lastname: Tilanus
mesh_terms: D000483:Alleles; D030561:Databases, Nucleic Acid; D005260:Female; D005805:Genes, MHC Class I / Q000235:genetics*; D006650:Histocompatibility Testing / Q000379:methods*; D006801:Humans; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods*; D012333:RNA, Messenger / Q000235:genetics*
chemical_list: D012333:RNA, Messenger
keywords: HLA class I; RNA sequence based typing; full length human leukocyte antigen gene polymorphism