HIGH-RESOLUTION HLA TYPING FOR THE DQB1 GENE BY SEQUENCE-BASED TYPING

The ideal high-resolution typing strategy for polymorphic genes is seq uence-based tl;ping. SET of genomic DNA has been developed for the HLA class Il gene DRB1, DRB3/4/5 and DPB1. For the DQB1 gene the sequence -based typing method was shown to cause a number of problems. To resol ve those problems, different primers to amplify and sequence exon 2 of DQB1 were designed and tested. With several printer combinations, pre ferential amplification was obsewed in individuals heterozygous for DQ B102/*03 and DQB1*02/*04. The preference was for DQB1*02 in many inst ances but could also be demonstrated for DQB103 or *04 and resulted o ccasionally in allelic dropout. The best primer combination was select ed and successfully used to type individuals heterozygous for DQB102, 03 and *04. To distinguish DQB1*0201 and *0202 primers for amplifica tion and sequencing of exon 3 were developed and correct subtyping was obtained. The ambiguous typing DQB10301/*0302 and DQB1*0303/*0304 wa s resolved bu allele-specific amplification and sequencing. A total of 258 individuals were fully typed for their DQB1 subtypes. All samples had been previously typed by PCR-SSP and serology. Concordant typing results were obtained for all individuals tested. The DQB1 alleles det ected included 0501, *0502, *0503, *0601, *0602, *0603, *0604, *0609, 0201, *0202, *0301, *0302. *0303, *0304, *0401 and *0402. Sequence-b ased typing of the DQB1 gene proved a reliable typing strategy for ass ignment of the different DQB1 alleles alter intensive selection of pri mers and test conditions.