OR47 The mystery of the missing DRB1 allele – solved by NGS!!!

The mystery began when HLA typing, of two apparently unrelated individuals at two different institutions, identified the presence of a DRB5∗01 gene without any evidence of an expected linked DRB1∗15, DRB1∗16, or, in rare cases, DRB1∗01 allele. Historic data from Sanger sequence-based typing included DRB1∗14, DRB3∗02:02, DRB5∗01:01, DQB1∗05:03, 06:02 for Individual 1 and DRB1∗04:01; DRB4∗01; DRB5∗01:01; DQB1∗03:01, 06:02 for Individual 2. Since those analyses only assessed exon 2, the apparent absence of a DRB1 allele associated with the DRB5 gene raised the possibility of an entire linked DRB1 gene having gone missing. Both specimens were recently re-examined using next generation sequencing technology. HLA-A, B, C genes were assessed for exons 1–7; DRB1, DRB3/B4/B5, DQA1, DQB1, and DPB1 were assessed for exons 1–4; and DPA1 for exons 2–4. In Individual 1, NGS identified DRB1∗14:54:01 and a DRB1∗15 exon 1 sequence, but without any evidence of DRB1∗15 sequences for exons 2 and 3. Since the exon 4 sequences of DRB1∗14:54:01 and DRB1∗15 are identical, it could not be determined whether the DRB1∗15 exon 4 sequence was present. Individual 2, typed as DRB1∗04:01:01 and, again, a DRB1∗15 exon 1. In the latter case, it was clear that no exon 2, 3, or 4 sequences were present for the DRB1∗15 gene, since exon 4 sequences differ between DRB1∗04:01:01 and DRB1∗15. The table shows the full NGS typing data. The HLA alleles shared by both individuals suggest a putative extended haplotype: A∗25:01:01-C∗12:03:01-B∗18:01:01-DRB1∗15new-DRB5∗01:01:01-DQA1∗01:02:01-DQB1∗06:02:01-DPA1∗01:03:01-DPB1∗23:01:01. NGS typing solved the missing allele mystery through identification of a truncated DRB1∗15 allele comprising an exon 1 sequence but without exons 2, 3, or 4. Similarly, there is DRB4∗03:01N which has an exon 3 sequence of a DRB4∗01:01, but without an exon 2 sequence. These unusual alleles appear to be the results of historic multi-exon deletion events and add to the astounding diversity of the human major histocompatibility complex. Download high-res image (242KB) Download full-size image A.G. Smith: 2. Consultant; Company/Organization; Scisco Genetics Inc. S.E. Pereira: 4. Scientific/Medical Advisor; Company/Organization; Scisco Genetics Inc. C. Pyo: 4. Scientific/Medical Advisor; Company/Organization; Scisco Genetics Inc. W. Nelson: 5. Employee; Company/Organization; Scisco Genetics Inc. M.Z. Askar: 3. Speaker’s Bureau; Company/Organization; Immucor. 4. Scientific/Medical Advisor; Company/Organization; Illumina. D.E. Geraghty: 6. Stock Shareholder; Company/Organization; Scisco Genetics Inc.