ArticleBMC genomics2017
HLA genotyping by next-generation sequencing of complementary DNA.
Article in BMC genomics, 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 6 papers.
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Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
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Who cites it
6 citing papers in PubMed.
- Present-day Admixed Genomes Reveal Prehistoric Adaptation to Cold, Maritime Diet, and Local Pathogens in Patagonia.Genome biology and evolution · 2026Article
- Article
- Simultaneous genotyping for human platelet antigen systems andFrontiers in immunology · 2022Article
- Capturing Differential Allele-Level Expression and Genotypes of All Classical HLA Loci and Haplotypes by a New Capture RNA-Seq Method.Frontiers in immunology · 2020Article
- Bioinformatics Strategies, Challenges, and Opportunities for Next Generation Sequencing-Based HLA Genotyping.Transfusion medicine and hemotherapy : offizielles Organ der Deutschen Gesellschaft fur Transfusionsmedizin und Immunhamatologie · 2019Review
- Adapting Cord Blood Collection and Banking Standard Operating Procedures for HLA-Homozygous Induced Pluripotent Stem Cells Production and Banking for Clinical Application.Journal of clinical medicine · 2019Review
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Authors and funding
3 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundGenotyping of the human leucocyte antigen (HLA) is indispensable for various medical treatments. However, unambiguous genotyping is technically challenging due to high polymorphism of the corresponding genomic region. Next-generation sequencing is changing the landscape of genotyping. In addition to high throughput of data, its additional advantage is that DNA templates are derived from single molecules, which is a strong merit for the phasing problem. Although most currently developed technologies use genomic DNA, use of cDNA could enable genotyping with reduced costs in data production and analysis. We thus developed an HLA genotyping system based on next-generation sequencing of cDNA.
methodsEach HLA gene was divided into 3 or 4 target regions subjected to PCR amplification and subsequent sequencing with Ion Torrent PGM. The sequence data were then subjected to an automated analysis. The principle of the analysis was to construct candidate sequences generated from all possible combinations of variable bases and arrange them in decreasing order of the number of reads. Upon collecting candidate sequences from all target regions, 2 haplotypes were usually assigned. Cases not assigned 2 haplotypes were forwarded to 4 additional processes: selection of candidate sequences applying more stringent criteria, removal of artificial haplotypes, selection of candidate sequences with a relaxed threshold for sequence matching, and countermeasure for incomplete sequences in the HLA database.
resultsThe genotyping system was evaluated using 30 samples; the overall accuracy was 97.0% at the field 3 level and 98.3% at the G group level. With one sample, genotyping of DPB1 was not completed due to short read size. We then developed a method for complete sequencing of individual molecules of the DPB1 gene, using the molecular barcode technology.
conclusionThe performance of the automatic genotyping system was comparable to that of systems developed in previous studies. Thus, next-generation sequencing of cDNA is a viable option for HLA genotyping.
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