More than 150 novel variants of HLA class I genes detected in German Stem Cell Donor Registry and UCLA International Cell Exchange samples

Abstract

High throughput analysis using amplicon-based next-generation sequencing (NGS) of HLA class I genes in samples of registered stem cell donors of the German Stem Cell Donor Registry Düsseldorf revealed 151 novel variants. In addition, four new variants were identified in well-defined samples obtained from the UCLA International Cell Exchange program. New alleles included 37 HLA-A, 57 HLA-B, and 61 HLA-C variant alleles. All variants were confirmed by NGS of HLA-A, HLA-B, and HLA-C genes including the respective 5' and 3' untranslated regions as well as Sanger sequence analysis. Mainly, the variants encompass single nucleotide changes in intronic as well as exonic parts of the genes. We identified intronic variations in 114 new alleles, nonsynonymous nucleotide changes in 25 alleles, synonymous nucleotide changes in nine alleles, and three hybrid alleles. Four alleles carry exonic deletions or insertions resulting in frameshift of peptide translation. Two novel alleles of HLA-C were shown to result in splicing defects of the transcript. Two alleles showed exonic as well as intronic changes. Thirty-four of the new alleles were found in multiple samples.

Nomenclature for factors of the HLA system, update February 2018

“…Additional information pertaining to new sequences is often included in the publications describing these alleles; a listing of recent publications that describe new HLA sequences is given in Table . An additional 155 alleles were recently published but have not been included in Table 3 due to space considerations . All new and confirmatory sequences should now be submitted directly to the World Health Organization (WHO) Nomenclature Committee for Factors of the HLA System via the IPD‐IMGT/HLA Database using the sequence submission tool provided .…”

“…An additional 155 alleles were recently published but have not been included in Table 3 due to space considerations. 2 All new and confirmatory sequences should now be submitted directly to the World Health Organization (WHO) Nomenclature Committee for Factors of the HLA System via the IPD-IMGT/HLA Database using the sequence submission tool provided. 3 The IPD-IMGT/HLA Database may be accessed via the WorldWide Web at: www.ebi.ac.uk/ipd/imgt/hla.…”

A novel HLA‐B variant, HLA‐B*14:02:16, discovered by amplicon‐based next‐generation sequencing

“…We used an amplicon‐based approach to analyse samples of newly registered potential stem cell donors …”

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We used an amplicon-based approach to analyse samples of newly registered potential stem cell donors. 1 Briefly, amplification of exons 2, 3, and 4 of HLA class I and HLA-DPB1 genes, and exons 2 and 3 of HLA-DRB1 and HLA-DQB1 (in parallel with selected exons of the blood group system genes ABO and Rh) was carried out in six multiplex polymerase chain reactions (PCRs). After a purification step using paramagnetic beads, amplificates underwent a second-round PCR that adds sample-specific barcodes and Illumina compatible adapter sequences.

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A novel HLA‐A null allele, HLA‐A*30:130N, discovered by amplicon‐based next‐generation sequencing

“…We used an amplicon-based approach to analyse HLAclass I and HLA-class II genes (in parallel with selected exons of the blood group system genes ABO and Rh). 1 Briefly, amplification of exons 2, 3, and 4 of HLA class I and HLA-DPB1 genes, and exons 2 and 3 of HLA-DRB1 and HLA-DQB1 were carried out in six multiplex polymerase chain reactions (PCRs). After a purification step using paramagnetic beads, amplificates underwent a second-round PCR that adds sample-specific barcodes and Illumina compatible adapter sequences.…”