Typing for all known MICA alleles by group-specific PCR and SSOP

Major histocompatibility complex class I chain-related gene (MICA) is a recently discovered polymorphic gene in the HLA region expressed mainly by certain epithelial cells, keratinocytes, endothelial cells, fibroblasts, and monocytes. MICA is structurally quite different from the HLA class I genes and is potentially associated with some diseases and with immune response to transplants. Some DNA-based typing techniques have previously been described for MICA including sequence-based typing (SBT) and analysis of single strand conformational polymorphisms (SSCP). In the present experiments we have developed a strategy that allows identification of all 54 MICA alleles described so far, using group-specific polymerase chain reactions (PCR) and sequence-specific oligonucleotide probes (SSOP). To analyze for the polymorphisms in exons 2, 3, and 4 an initial screening with group-specific primers, based on polymorphism at position 69 of exon 2, and at position 615-616 of exon 4, was used to determine four major groups of alleles. Then group-specific PCR amplifications were performed and the amplified DNA was hybridized with the corresponding panels of SSOP. An additional amplification was performed with locus-specific primers and hybridized with a set of SSOP to identify and/or confirm the presence of some of the alleles. Unequivocal MICA typing was achieved for 97 of 103 individuals. Of 54 previously described alleles, only 14 were observed in this population. One unexpected hybridization pattern was observed, and molecular cloning and sequencing confirmed it to be a novel sequence, which was given the local designation MICA-055D. The gene frequencies among 103 unrelated North American Caucasian donors were determined and the linkage disequilibrium between MICA and HLA-B was analyzed.