Performance Characteristics and Validation of Next-Generation Sequencing for Human Leucocyte Antigen Typing

Weimer, Eric T.; Montgomery, Maureen C.; Petraroia, Rosanne; Crawford, John; Schmitz, John L.

doi:10.1016/j.jmoldx.2016.03.009

Cited by 60 publications

(40 citation statements)

References 24 publications

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“…These were easily detected due to non‐common linkage disequilibrium. Most studies have reported very low allele dropout frequencies (<1%), whereas others have reported frequencies as high as 10%, especially for the HLA‐DQB1 locus . Therefore, the need for alternative methods to confirm apparently homozygous HLA alleles is a matter of debate.…”

Section: Discussionmentioning

confidence: 99%

“…The use of multiplex identifiers allows samples to be tagged and pooled, increasing throughput and decreasing costs. One of the first platforms used for HLA typing was the 454/GS FLX sequencer (Roche), [13][14][15] which has been progressively supplanted by Illumina [16][17][18][19][20] and Ion Torrent technologies. [21][22][23][24] Advanced technologies are also used, such as Pacific Biosciences's single-molecule real-time sequencing [25][26][27][28][29][30] and Oxford Nanopore single-cell DNA template strand sequencing.…”

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confidence: 99%

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Evaluation of the AllType kit for HLA typing using the Ion Torrent S5 XL platform

Cargou

Ralazamahaleo

Blouin

et al. 2019

HLA

147

141

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HLA genotyping by next‐generation sequencing is now widely performed. We aimed at evaluating the performance of the One Lambda AllType kit using Thermo Fisher Scientific reagents on the Ion S5 XL platform. Reads were analyzed using the TypeStream Visual software. We performed 15 runs between April and September 2018 to type DNA at the HLA‐A/B/C/DRB1/3/4/5/DQA1/DQB1/DPA1/DPB1 loci from 340 samples and 15 positive controls. We observed only seven (0.1%) critical mistakes among the 6009 alleles typed, corresponding to two allele dropouts, one false heterozygous typing assignment, and four phasing abnormalities. Among the 1793 presumably new alleles detected by the analysis software, 11 displayed exon mismatches, of which nine were confirmed as new alleles and two had been described previously. Intron mismatches were observed among the remaining presumably new alleles, of which 371 were considered as probably new, and 1411 were rejected for at least one sequence feature such as homopolymers (n = 1206), nucleotide doublet repeats (n = 26), low read depth (<200 reads, n = 93), high background (>20%, n = 79), or phasing abnormalities (n = 7). A comparison of the AllType results with those obtained using other methods at the second‐field resolution level showed 99.5% (1497/1504) concordance for the HLA‐A/B/C/DRB1/DQB1/DPB1 loci. Similar agreement was observed between the HLA‐C or HLA‐DRB3/4/5 results and common linkage disequilibrium, with 96.6% (657/680) and 97.2% (530/545) concordance, respectively. Therefore, the AllType kit used with the Ion S5 XL platform displayed satisfactory performance for HLA typing in current clinical practice.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Evaluation of the AllType kit for HLA typing using the Ion Torrent S5 XL platform

Cargou

Ralazamahaleo

Blouin

et al. 2019

HLA

147

141

View full text Add to dashboard Cite

show abstract

“…One new emerging technology that can be applied to achieve HLA typing is next generation sequencing (NGS) [12], [49], [50]. Such technologies enabling single molecule sequencing in high throughput are becoming more widely available for HLA typing with more accurate computational tools [51] and implementation in clinical HLA laboratories [52], [53]. However, accurate phasing is reliant on generating large enough fragment sizes which is currently limiting.…”

Section: Discussionmentioning

confidence: 99%

High resolution HLA haplotyping by imputation for a British population bioresource

et al. 2017

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This study aimed to establish the occurrence and frequency of HLA alleles and haplotypes for a healthy British Caucasian population bioresource from Oxfordshire. We present the results of imputation from HLA SNP genotyping data using SNP2HLA for 5553 individuals from Oxford Biobank, defining one- and two-field alleles together with amino acid polymorphisms. We show that this achieves a high level of accuracy with validation using sequence-specific primer amplification PCR. We define six- and eight-locus HLA haplotypes for this population by Bayesian methods implemented using PHASE. We determine patterns of linkage disequilibrium and recombination for these individuals involving classical HLA loci and show how analysis within a haplotype block structure may be more tractable for imputed data. Our findings contribute to knowledge of HLA diversity in healthy populations and further validate future large-scale use of HLA imputation as an informative approach in population bioresources.

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“…First, unambiguous genotypes are generated as a result of contiguous full‐length sequence amplifications from the 5′UTR through the 3′UTR regions. Several NGS HLA typing methods for class II loci focus on core exons 2 to 3 and omit exon 1 and/or portions of other exons 5,7,20‐24 . These approaches can lead to ambiguous calls and reduce the likelihood of identification of novel or rare alleles.…”

Section: Discussionmentioning

confidence: 99%

High‐throughput next‐generation sequencing to genotype six classical HLA loci from 96 donors in a single MiSeq run

Ehrenberg

Geretz

Sindhu

et al. 2017

HLA

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Next generation sequencing (NGS) methods have been established as an efficient approach for HLA typing because unlike traditional Sanger sequencing, they provide unambiguous results at a reasonable cost. We previously developed a multi-locus index method to genotype four HLA loci (A, B, C, and DRB1) on the Illumina MiSeq platform. We have now expanded this method to include two additional loci, HLA-DPB1 and DQB1. Contiguous full-length amplicons from 5'UTR through 3'UTR regions were generated using one long-range PCR reaction per locus for each of the six loci from 96 individuals of different ethnicities. The six amplicons from each donor were pooled, enzymatically fragmented and given a donor-specific index. This approach enabled sequencing of 576 loci from 96 individuals in a single MiSeq run. Donor-specific sequence reads were demultiplexed, and allele calls were generated from FASTQ files using commercially available software. Comparison to HLA genotypes generated from Sanger sequence-based typing (SBT) identified no discordances among any of the alleles analyzed in this study. Importantly, this method was able to resolve 22 DPB1 and 20 DQB1 alleles that were ambiguous with the SBT method. Furthermore, a novel allele in each of these two loci was identified, with the DQB1*05:01:24 allele having a frequency of greater than five percent. This method was subsequently validated against a blinded panel of 22 samples from the 17th International HLA and Immunogenetics Workshop. The flexibility of the method is further highlighted by successful genotyping of eight loci comprising all classical HLA loci for a subset of the samples. We now present a high-throughput, high-resolution, scalable NGS HLA typing method to accurately and efficiently genotype all classical HLA class I and II loci.

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Performance Characteristics and Validation of Next-Generation Sequencing for Human Leucocyte Antigen Typing

Cited by 60 publications

References 24 publications

Evaluation of the AllType kit for HLA typing using the Ion Torrent S5 XL platform

Evaluation of the AllType kit for HLA typing using the Ion Torrent S5 XL platform

High resolution HLA haplotyping by imputation for a British population bioresource

High‐throughput next‐generation sequencing to genotype six classical HLA loci from 96 donors in a single MiSeq run

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