Identification of two major HLA-B44 subtypes and a novel B44 sequence (B*4404). Oligotyping and solid phase sequencing of polymerase chain reaction products.
PCR-based analyses were performed for the identification of HLA-B44 subgroups. Genomic DNA from six homozygous cell lines and 44 healthy individuals who had serologically tested positive for HLA-B44 was investigated for polymorphism in exons 2 and 3 of the HLA-B44 genes. Two primers were designed for specific amplification of the B*4401 allele in exon 2. None of the tested genomic DNAs, including the cell line "BAU-J" from which the sequence of B*4401 was derived, was amplified successfully using these primers, indicating that the B*4401 sequence may not be correct in position 242-244. For identification of the B*4402 and *4403 subtypes we specifically amplified the B44 gene in exon 3 using two sequence-specific primers. The PCR products, which were obtained from all B44-positive samples (n = 50) and from none of the B44-negative controls (n = 20), were subsequently hybridized with the dig-ddUTP-labeled oligonucleotides. The base substitution at position 146, as described previously for B*4401 and *4402 (C for G), could not be confirmed by oligonucleotide hybridization. In contrast, the oligonucleotide typing for G in position 146 gave positive signals in all B44-positive samples. Except for one, HLA-B44-positive DNAs from LCLs and healthy individuals could be divided into two subgroups according to the polymorphic region in position 195-197. Out of 44 unrelated individuals with B44, 27 (61%) were positive for B*4402 and 16 (36%) were positive for B*4403.(ABSTRACT TRUNCATED AT 250 WORDS)
Yao
Z
Z
Immunogenetics Laboratory, University of Munich, Germany.
Keller
E
E
Scholz
S
S
McNicholas
A
A
Volgger
A
A
Albert
E D
ED
D000483:Alleles; D001483:Base Sequence; D002460:Cell Line; D016384:Consensus Sequence; D005091:Exons / Q000235:genetics; D005240:Feasibility Studies; D015235:HLA-B Antigens / Q000145:classification / Q000235:genetics*; D059948:HLA-B44 Antigen; D006650:Histocompatibility Testing / Q000379:methods*; D006801:Humans; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid
D015235:HLA-B Antigens; D059948:HLA-B44 Antigen; D015345:Oligonucleotide Probes