AUTHOR=Anukul Nampeung , Jenjaroenpun Piroon , Sirikul Chonticha , Wankaew Natnicha , Nimsamer Pattaraporn , Roothumnong Ekkapong , Pithukpakorn Manop , Leetrakool Nipapan , Wongsurawat Thidathip 

TITLE=Ultrarapid and high-resolution HLA class I typing using transposase-based nanopore sequencing applied in pharmacogenetic testing

JOURNAL=Frontiers in Genetics

VOLUME=14

YEAR=2023

URL=https://www.frontiersin.org/journals/genetics/articles/10.3389/fgene.2023.1213457

DOI=10.3389/fgene.2023.1213457

ISSN=1664-8021

ABSTRACT=<p>Nanopore sequencing has been examined as a method for rapid and high-resolution human leukocyte antigen (HLA) typing in recent years. We aimed to apply ultrarapid nanopore-based HLA typing for HLA class I alleles associated with drug hypersensitivity, including HLA-A*31:01, HLA-B*15:0<italic>2</italic>, and HLA-C*08:01. Most studies have used the Oxford Nanopore Ligation Sequencing kit for HLA typing, which requires several enzymatic reactions and remains relatively expensive, even when the samples are multiplexed. Here, we used the Oxford Nanopore Rapid Barcoding kit, which is transposase-based, with library preparation taking less than 1 h of hands-on time and requiring minimal reagents. Twenty DNA samples were genotyped for HLA-A, -B, and -C; 11 samples were from individuals of different ethnicity and nine were from Thai individuals. Two primer sets, a commercial set and a published set, were used to amplify the <italic>HLA-A</italic>, -<italic>B</italic>, and -<italic>C</italic> genes. HLA-typing tools that used different algorithms were applied and compared. We found that without using several third-party reagents, the transposase-based method reduced the hands-on time from approximately 9 h to 4 h, making this a viable approach for obtaining same-day results from 2 to 24 samples. However, an imbalance in the PCR amplification of different haplotypes could affect the accuracy of typing results. This work demonstrates the ability of transposase-based sequencing to report 3-field HLA alleles and its potential for race- and population-independent testing at considerably decreased time and cost.</p>