{"title":"两步PCR-SSP和纳米孔测序对HLA-B*57:01和HLA-B*58:01分型的一致性比较研究","authors":"Nattapong Intharuangrung, Chonticha Sirikul, Pattaraporn Nimsamer, Thidathip Wongsurawat, Auttachai Saejia, Nampeung Anukul","doi":"10.3389/fgene.2025.1649990","DOIUrl":null,"url":null,"abstract":"<p><p>This study reports an optimized in-house 2-step PCR-SSP assay for rapid, cost-effective detection of <i>HLA-B*57:01</i> and <i>HLA-B*58:01</i> in routine pharmacogenomics laboratory. This assay employs allele-specific primers positioned within exon 2-3 boundaries, validated <i>in silico</i> against common <i>HLA-B</i> alleles. Using 30 clinical DNA samples, our PCR workflow (<1 h) showed 100% concordance at 2-field resolution with Oxford Nanopore sequencing performed using ligation-based sequencing kit with PCR barcoding. Cohen's kappa was 1.00 with 95% CI. The turnaround time and reagent cost per sample were reduced to 1 h of hands-on PCR time and USD 7 per sample, respectively. These do not include DNA extraction or gel electrophoresis analysis. This 2-step PCR-SSP offers a robust alternative for pharmacogenomic screening in resource-limited settings for detecting the <i>HLA-B*57:01</i> and <i>HLA-B*58:01</i>.</p>","PeriodicalId":12750,"journal":{"name":"Frontiers in Genetics","volume":"16 ","pages":"1649990"},"PeriodicalIF":2.8000,"publicationDate":"2025-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12507366/pdf/","citationCount":"0","resultStr":"{\"title\":\"Concordance of an in-house 2-steps PCR-SSP and nanopore sequencing for <i>HLA-B*57:01</i> and <i>HLA-B*58:01</i> typing: a comparative study.\",\"authors\":\"Nattapong Intharuangrung, Chonticha Sirikul, Pattaraporn Nimsamer, Thidathip Wongsurawat, Auttachai Saejia, Nampeung Anukul\",\"doi\":\"10.3389/fgene.2025.1649990\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>This study reports an optimized in-house 2-step PCR-SSP assay for rapid, cost-effective detection of <i>HLA-B*57:01</i> and <i>HLA-B*58:01</i> in routine pharmacogenomics laboratory. This assay employs allele-specific primers positioned within exon 2-3 boundaries, validated <i>in silico</i> against common <i>HLA-B</i> alleles. Using 30 clinical DNA samples, our PCR workflow (<1 h) showed 100% concordance at 2-field resolution with Oxford Nanopore sequencing performed using ligation-based sequencing kit with PCR barcoding. Cohen's kappa was 1.00 with 95% CI. The turnaround time and reagent cost per sample were reduced to 1 h of hands-on PCR time and USD 7 per sample, respectively. These do not include DNA extraction or gel electrophoresis analysis. This 2-step PCR-SSP offers a robust alternative for pharmacogenomic screening in resource-limited settings for detecting the <i>HLA-B*57:01</i> and <i>HLA-B*58:01</i>.</p>\",\"PeriodicalId\":12750,\"journal\":{\"name\":\"Frontiers in Genetics\",\"volume\":\"16 \",\"pages\":\"1649990\"},\"PeriodicalIF\":2.8000,\"publicationDate\":\"2025-09-25\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12507366/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Frontiers in Genetics\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.3389/fgene.2025.1649990\",\"RegionNum\":3,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2025/1/1 0:00:00\",\"PubModel\":\"eCollection\",\"JCR\":\"Q2\",\"JCRName\":\"GENETICS & HEREDITY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Frontiers in Genetics","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.3389/fgene.2025.1649990","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2025/1/1 0:00:00","PubModel":"eCollection","JCR":"Q2","JCRName":"GENETICS & HEREDITY","Score":null,"Total":0}
Concordance of an in-house 2-steps PCR-SSP and nanopore sequencing for HLA-B*57:01 and HLA-B*58:01 typing: a comparative study.
This study reports an optimized in-house 2-step PCR-SSP assay for rapid, cost-effective detection of HLA-B*57:01 and HLA-B*58:01 in routine pharmacogenomics laboratory. This assay employs allele-specific primers positioned within exon 2-3 boundaries, validated in silico against common HLA-B alleles. Using 30 clinical DNA samples, our PCR workflow (<1 h) showed 100% concordance at 2-field resolution with Oxford Nanopore sequencing performed using ligation-based sequencing kit with PCR barcoding. Cohen's kappa was 1.00 with 95% CI. The turnaround time and reagent cost per sample were reduced to 1 h of hands-on PCR time and USD 7 per sample, respectively. These do not include DNA extraction or gel electrophoresis analysis. This 2-step PCR-SSP offers a robust alternative for pharmacogenomic screening in resource-limited settings for detecting the HLA-B*57:01 and HLA-B*58:01.
Frontiers in GeneticsBiochemistry, Genetics and Molecular Biology-Molecular Medicine
CiteScore
5.50
自引率
8.10%
发文量
3491
审稿时长
14 weeks
期刊介绍：
Frontiers in Genetics publishes rigorously peer-reviewed research on genes and genomes relating to all the domains of life, from humans to plants to livestock and other model organisms. Led by an outstanding Editorial Board of the world’s leading experts, this multidisciplinary, open-access journal is at the forefront of communicating cutting-edge research to researchers, academics, clinicians, policy makers and the public.
The study of inheritance and the impact of the genome on various biological processes is well documented. However, the majority of discoveries are still to come. A new era is seeing major developments in the function and variability of the genome, the use of genetic and genomic tools and the analysis of the genetic basis of various biological phenomena.
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