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Development of a high-resolution NGS-based HLA-typing and analysis pipeline

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Item Type:Article
Title:Development of a high-resolution NGS-based HLA-typing and analysis pipeline
Creators Name:Wittig, M. and Anmarkrud, J.A. and Kässens, J.C. and Koch, S. and Forster, M. and Ellinghaus, E. and Hov, J.R. and Sauer, S. and Schimmler, M. and Ziemann, M. and Görg, S. and Jacob, F. and Karlsen, T.H. and Franke, A.
Abstract:The human leukocyte antigen (HLA) complex contains the most polymorphic genes in the human genome. The classical HLA class I and II genes define the specificity of adaptive immune responses. Genetic variation at the HLA genes is associated with susceptibility to autoimmune and infectious diseases and plays a major role in transplantation medicine and immunology. Currently, the HLA genes are characterized using Sanger- or next-generation sequencing (NGS) of a limited amplicon repertoire or labeled oligonucleotides for allele-specific sequences. High-quality NGS-based methods are in proprietary use and not publicly available. Here, we introduce the first highly automated open-kit/open-source HLA-typing method for NGS. The method employs in-solution targeted capturing of the classical class I (HLA-A, HLA-B, HLA-C) and class II HLA genes (HLA-DRB1, HLA-DQA1, HLA-DQB1, HLA-DPA1, HLA-DPB1). The calling algorithm allows for highly confident allele-calling to three-field resolution (cDNA nucleotide variants). The method was validated on 357 commercially available DNA samples with known HLA alleles obtained by classical typing. Our results showed on average an accurate allele call rate of 0.99 in a fully automated manner, identifying also errors in the reference data. Finally, our method provides the flexibility to add further enrichment target regions.
Keywords:Alleles, DNA Sequence Analysis, HLA Antigens, High-Throughput Nucleotide Sequencing, Histocompatibility Testing, Software
Source:Nucleic Acids Research
ISSN:0305-1048
Publisher:Oxford University Press (U.K.)
Volume:43
Number:11
Page Range:e70
Date:23 June 2015
Official Publication:https://doi.org/10.1093/nar/gkv184
PubMed:View item in PubMed

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