HLA-CW allele analysis by PCR-restriction fragment length polymorphism: Study of known and additional alleles

Zohreh Tatari, Catherine Fortier, Vlasta Bobrynina, Pascale Loiseau, Dominique Charron, Colette Raffoux

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Abstract

We describe a technique for HLA-Cw genotyping by digestion of PCR- amplified genes with restriction endonucleases. Locus-specific primers selectively amplified HLA-Cw sequences from exon 2 in a single PCR that avoided coamplification of other classical and nonclassical class I genes. Amplified DNAs were digested with selected enzymes. Sixty-three homozygous cell lines from International Histocompatibility Workshop X and 113 unrelated individual cells were genotyped for HLA-Cw and compared with serology. The present protocol can distinguish 23 alleles corresponding to the known HLA- Cw sequences. Genotyping of serologically undetectable alleles (HLA-Cw Blank) and of heterozygous cells was made possible by using this method. Six additional HLA-Cw alleles were identified by unusual restriction patterns and confirmed by sequencing; this observation suggests the presence of another family of allele-sharing clusters in the HLA-B locus. This PCR-restriction endonuclease method provides a simple and convenient approach for HLA-Cw DNA typing, allowing the definition of serologically undetectable alleles, and will contribute to the evaluation of the biological role of the HLA-C locus.

Original languageEnglish
Pages (from-to)8803-8807
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume92
Issue number19
DOIs
Publication statusPublished - 12 Sep 1995

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