Sequence analysis of HLA class II domains: characterization of the DQw3 family of DQB genes.

HLA class II allelic variants within the DQw3-related family of genes carry distinct allo-specificities and have been implicated in specific HLA-disease associations, such as insulin-dependent diabetes mellitus. To investigate the nucleotide variations which characterize DQw3 genes, we applied a novel cDNA cloning strategy that uses a single-stranded vector/primer system to facilitate DNA sequencing of allelically variable gene families. Using a DQB-specific primer sequence and M13 bacteriophage as the cloning vector, direct cloning and sequencing of multiple DQB genes was performed without the need for second strand synthesis or for subcloning. Sequence analysis from eight lymphoblastoid cell lines selected to represent different ethnic backgrounds revealed three DQw3-related DQB genes, DQB3.1, 3.2, and 3.3, corresponding to the newly designated HLA-DQw7, w8, and w9 specificities, respectively. An unusual Pro-Pro couplet at codons 55-56 is characteristic of all DQw3-positive sequences and may be contributing to the broad DQw3 allospecificity. Comparisons among ethnically disparate DQw3-related sequences showed no additional expressed or silent nucleotide substitutions among these DQB alleles. Thus, polymorphism within the DQw3 family of genes appears to be extremely limited, with a paucity of nucleotide variations accumulated by evolutionary distance.