Allele and haplotype frequencies of human leukocyte antigen-A, -B, -C, -DRB1, -DRB3/4/5, -DQA1, -DQB1, -DPA1, and -DPB1 by next generation sequencing-based typing in Koreans in South Korea

Allele frequencies and haplotype frequencies of HLA-A, -B, -C, -DRB1, -DRB3/4/5, -DQA1, -DQB1, -DPA1, and -DPB1 have been rarely reported in South Koreans using unambiguous, phase-resolved next generation DNA sequencing. In this study, HLA typing of 11 loci in 173 healthy South Koreans were performed using next generation DNA sequencing with long-range PCR, TruSight® HLA v2 kit, Illumina MiSeqDx platform system, and Assign™ for TruSight™ HLA software. Haplotype frequencies were calculated using the PyPop software. Direct counting methods were used to investigate the association with DRB1 for samples with only one copy of a particular secondary DRB locus. We compared these allele types with the ambiguous allele combinations of the IPD-IMGT/HLA database. We identified 20, 40, 26, 31, 19, 16, 4, and 16 alleles of HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DQA1, HLA-DQB1, HLA-DPA1, and HLA-DPB1, respectively. The number of HLA-DRB3/4/5 alleles was 4, 5, and 3, respectively. The haplotype frequencies of most common haplotypes were as follows: A*33:03:01-B*44:03:01-C*14:03-DRB1*13:02:01-DQB1*06:04:01-DPB1*04:01:01 (2.89%), A*33:03:01-B*44:03:01-C*14:03 (4.91%), DRB1*08:03:02-DQA1*01:03:01-DQB1*06:01:01-DPA1*02:02:02-DPB1*05:01:01 (5.41%), DRB1*04:05:01-DRB4*01:03:01 (12.72%), DQA1*01:03:01-DQB1*06:01:01 (13.01%), and DPA1*02:02:02-DPB1*05:01:01 (30.83%). In samples with only one copy of a specific secondary DRB locus, we examined its association with DRB1. We, thus, resolved 10 allele ambiguities in HLA-B, -C (each exon 2+3), -DRB1, -DQB1, -DQA1, and -DPB1 (each exon 2) of the IPD-IMGT/HLA database. Korean population was geographically close to Japanese and Han Chinese populations in the genetic distances by multidimensional scaling (MDS) plots. The information obtained by HLA typing of the 11 extended loci by next generation sequencing may be useful for more exact diagnostic tests on various transplantations and the genetic population relationship studies in South Koreans.

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HLA‐A , ‐B, ‐C, ‐ DRB1 allele and haplotype frequencies of the Korean population and performance characteristics of HLA typing by next‐generation sequencing

HLA ◽

10.1111/tan.14167 ◽

2020 ◽

Author(s):

Dong Wook Jekarl ◽

Gun Dong Lee ◽

Jae Bin Yoo ◽

Jung Rok Kim ◽

Haein Yu ◽

...

Keyword(s):

Next Generation Sequencing ◽

Performance Characteristics ◽

Hla Typing ◽

Korean Population ◽

Next Generation ◽

Drb1 Allele ◽

Haplotype Frequencies ◽

And Performance ◽

Generation Sequencing

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Identification of DEEP Vein Thrombosis GENETIC RISK Variants by NEXT GENERATION Sequencing of Hemostatic Genes

Blood ◽

10.1182/blood.v118.21.710.710 ◽

2011 ◽

Vol 118 (21) ◽

pp. 710-710

Author(s):

Luca Andrea Lotta ◽

Mark Wang ◽

Jin Yu ◽

Fuli Yu ◽

Ida Martinelli ◽

...

Keyword(s):

Deep Vein Thrombosis ◽

Next Generation Sequencing ◽

Dna Sequencing ◽

Binomial Probability ◽

Next Generation ◽

Next Generation Dna Sequencing ◽

Vein Thrombosis ◽

Risk Variants ◽

Deep Vein ◽

Generation Sequencing

Abstract Abstract 710 Next-generation DNA sequencing is opening new avenues for genetic association studies in complex diseases that, like deep vein thrombosis (DVT), have a strong genetic basis only partially accounted for by currently identified risk variants. Using next-generation DNA sequencing, we resequenced the entire protein-coding area and intron-exon boundaries of ≈200 hemostatic/pro-inflammatory genes (including all coagulation factor and anticoagulant protein genes) in 26 Italian individuals with idiopathic DVT of the lower limbs and 28 age-, gender- and ethnicity-matched healthy controls. In order to enrich for genetic component, DVT patients were selected from a cohort of 730 cases of idiopathic DVT referred to the Angelo Bianchi Bonomi Hemophilia and Thrombosis Center (Milan, Italy) on the basis of the following criteria: (a) negative for anti-phospholipid autoantibodies, anticoagulant protein deficiencies, factor V Leiden and prothrombin G20210A, (b) age of onset <55 years. Genomic libraries with barcode sequence-tags were pooled (in pools of 8 or 16 samples) and enriched for target DNA sequences by capture on Roche NimbleGen HD2 2.1M-probe custom chips. Sequencing was performed on ABI SOLiD 4 platforms. After sequencing, reads with the barcodes were assigned to the corresponding sample and mapped to reference human genome, NCBI36/hg18, using BFAST software. We produced more than 18 gigabases of raw sequence data to sequence at high depth of coverage (>40X after duplicate read removal) the 700-kilobase target region. A total of 2351 single nucleotide variants (SNVs) and 121 short indels were identified. We developed a dedicated software in order to be able to run association analyses in our dataset using PLINK. A plain association analysis was used to compare the frequencies of common SNVs, whereas a gene-based analysis of restricted non-synonymous mutations was used to reveal potential associations of rare variants. Of 626 common SNVs tested for association, 32 SNVs from 13 genes were associated with p<0.01. These include 8 missense variants at 6 genes, 2 in genes already reported in association with DVT (FGA, ABO) and 6 in genes not previously reported (LPA, COL4A6, PLCG2 and KNG1). As a positive control we confirmed the association of rs6050 at FGA in ∼1400 Italian individuals (p=1.9 × 10−5, OR 1.45; 95% CI, 1.22–1.72). Replication of the other variants is ongoing. Gene-based analyses revealed an excess of private missense variants at anticoagulant protein genes (13 vs 3 variants; binomial probability, p=0.005) and at collagen 4 genes (14 vs 6 variants; binomial probability, p=0.02) in DVT cases compared to controls. We are currently expanding our investigations to 100 DVT cases and 100 controls. Although obtained in a relatively small number of patients and controls, our results illustrate the potential of next-generation sequencing for the discovery of genetic variation predisposing to complex diseases like DVT. Disclosures: No relevant conflicts of interest to declare.

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Targeted Next-Generation Sequencing for Human Leukocyte Antigen Typing in a Clinical Laboratory: Metrics of Relevance and Considerations for Its Successful Implementation

Archives of Pathology & Laboratory Medicine ◽

10.5858/arpa.2016-0537-ra ◽

2017 ◽

Vol 141 (6) ◽

pp. 806-812 ◽

Cited By ~ 19

Author(s):

Manish J. Gandhi ◽

Deborah Ferriola ◽

Yanping Huang ◽

Jamie L. Duke ◽

Dimitri Monos

Keyword(s):

Next Generation Sequencing ◽

Clinical Laboratory ◽

Human Leukocyte ◽

Hla Typing ◽

Successful Implementation ◽

Next Generation ◽

Human Leukocyte Antigen Typing ◽

Clinical Laboratories ◽

Exon 1 ◽

Generation Sequencing

Context.— Numerous feasibility studies to type human leukocyte antigens (HLAs) by next-generation sequencing (NGS) have led to the development of vendor-supported kits for HLA typing by NGS. Some clinical laboratories have introduced HLA-NGS, and many are investigating the introduction. Standards from accrediting agencies form the regulatory framework for introducing this test into clinical laboratories. Objectives.— To provide an assessment of metrics and considerations relevant to the successful implementation of clinical HLA-NGS typing, and to provide as a reference a validated HLA-NGS protocol used clinically since December 2013 at the Children's Hospital of Philadelphia (Philadelphia, Pennsylvania). Data Sources.— The HLA-NGS has been performed on 2532 samples. The initial 1046 and all homozygous samples were also typed by an alternate method. The HLA-NGS demonstrated 99.7% concordance with the alternate method. Ambiguous results were most common at the DPB1 locus because of a lack of phasing between exons 2 and 3 or the unsequenced exon 1 (533 of 2954 alleles; 18.04%) and the DRB1 locus because of not sequencing exon 1 (75 of 3972 alleles; 1.89%). No ambiguities were detected among the other loci. Except for 2 false homozygous samples, all homozygous samples (1891) demonstrated concordance with the alternate method. The article is organized to address the critical elements in the preanalytic, analytic, and postanalytic phases of introducing this assay into the clinical laboratory. Conclusions.— The results demonstrate that HLA typing by NGS is a highly accurate, reproducible, efficient method that provides more-complete sequencing information for the length of the HLA gene and can be the single methodology for HLA typing in clinical immunogenetics laboratories.

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Benchmarking the Human Leukocyte Antigen Typing Performance of Three Assays and Seven Next-Generation Sequencing-Based Algorithms

Frontiers in Immunology ◽

10.3389/fimmu.2021.652258 ◽

2021 ◽

Vol 12 ◽

Author(s):

Ping Liu ◽

Minya Yao ◽

Yu Gong ◽

Yunjie Song ◽

Yanan Chen ◽

...

Keyword(s):

Clinical Practice ◽

Next Generation Sequencing ◽

Human Leukocyte Antigen ◽

Human Leukocyte ◽

Hla Typing ◽

Read Length ◽

Next Generation ◽

Leukocyte Antigen ◽

The Third ◽

Generation Sequencing

With the great progress made recently in next generation sequencing (NGS) technology, sequencing accuracy and throughput have increased, while the cost for data has decreased. Various human leukocyte antigen (HLA) typing algorithms and assays have been developed and have begun to be used in clinical practice. In this study, we compared the HLA typing performance of three HLA assays and seven NGS-based HLA algorithms and assessed the impact of sequencing depth and length on HLA typing accuracy based on 24 benchmarked samples. The algorithms HISAT-genotype and HLA-HD showed the highest accuracy at both the first field and the second field resolution, followed by HLAscan. Our internal capture-based HLA assay showed comparable performance with whole exome sequencing (WES). We found that the minimal depth was 100X for HISAT-genotype and HLA-HD to obtain more than 90% accuracy at the third field level. The top three algorithms were quite robust to the change of read length. Thus, we recommend using HISAT-genotype and HLA-HD for NGS-based HLA genotyping because of their higher accuracy and robustness to read length. We propose that a minimal sequence depth for obtaining more than 90% HLA typing accuracy at the third field level is 100X. Besides, targeting capture-based NGS HLA typing may be more suitable than WES in clinical practice due to its lower sequencing cost and higher HLA sequencing depth.

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P262 Dna sequencing with Scisco genetics next generation sequencing HLA typing version 5 kit

Human Immunology ◽

10.1016/j.humimm.2017.06.322 ◽

2017 ◽

Vol 78 ◽

pp. 246

Author(s):

Arisa Oki ◽

Randa Abou-Taleb ◽

Jean F. Garcia-Gomez ◽

Ketevan Gendzekhadze ◽

David Senitzer

Keyword(s):

Next Generation Sequencing ◽

Dna Sequencing ◽

Hla Typing ◽

Next Generation ◽

Generation Sequencing

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Next-Generation Sequencing: From Basic Research to Diagnostics

Clinical Chemistry ◽

10.1373/clinchem.2008.112789 ◽

2009 ◽

Vol 55 (4) ◽

pp. 641-658 ◽

Cited By ~ 459

Author(s):

Karl V Voelkerding ◽

Shale A Dames ◽

Jacob D Durtschi

Keyword(s):

Next Generation Sequencing ◽

Dna Sequencing ◽

Single Molecule ◽

Basic Research ◽

Clinical Diagnostics ◽

Massively Parallel ◽

Next Generation ◽

New Paradigm ◽

The Impact ◽

Generation Sequencing

Abstract Background: For the past 30 years, the Sanger method has been the dominant approach and gold standard for DNA sequencing. The commercial launch of the first massively parallel pyrosequencing platform in 2005 ushered in the new era of high-throughput genomic analysis now referred to as next-generation sequencing (NGS). Content: This review describes fundamental principles of commercially available NGS platforms. Although the platforms differ in their engineering configurations and sequencing chemistries, they share a technical paradigm in that sequencing of spatially separated, clonally amplified DNA templates or single DNA molecules is performed in a flow cell in a massively parallel manner. Through iterative cycles of polymerase-mediated nucleotide extensions or, in one approach, through successive oligonucleotide ligations, sequence outputs in the range of hundreds of megabases to gigabases are now obtained routinely. Highlighted in this review are the impact of NGS on basic research, bioinformatics considerations, and translation of this technology into clinical diagnostics. Also presented is a view into future technologies, including real-time single-molecule DNA sequencing and nanopore-based sequencing. Summary: In the relatively short time frame since 2005, NGS has fundamentally altered genomics research and allowed investigators to conduct experiments that were previously not technically feasible or affordable. The various technologies that constitute this new paradigm continue to evolve, and further improvements in technology robustness and process streamlining will pave the path for translation into clinical diagnostics.

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