DNA polymorphism of the HLA-B35 gene associated to different HLA-C locus alleles

Peripheral blood leukocyte DNA from 31 donors, previously typed as HLA-B35, was digested with EcoRV and analyzed by Southern transfer and hybridization to an HLA class I probe. Out of 31 HLA-B35 positive (+) individuals, 24 showed a 4.6-kb band, previously associated with the B35 allele by Cohen et al. (Proc Natl Acad Sci USA 80:6289-6292). The HLA-B35 allele has a strong linkage disequilibrium (70%) with the Cw4 allele. All samples having the 4.6-kb band corresponded to HLA-B35+, Cw4+ cells, whereas those lacking the band came from HLA-B35+, Cw4-negative (-) cells. Five HLA-B35-, Cw4+ samples were also studied and none showed the band. An HLA-B locus-specific probe gave a strong hybridization signal to this fragment, whereas an HLA-C locus-specific probe revealed different bands. The results suggest that the 4.6-kb fragment contains the B35 gene. Digestion with other restriction enzymes, located the polymorphic site to the 3' end of the gene. Analysis of 18 additional individuals with other specificities of the "4c" antigenic cluster (HLA-B18, B51, B52 and B53) showed that the EcoRV/4.6 kb-band was also present in 5/5 B52 cells and 4/4 B53 samples.