HLA typing by next‐generation sequencing – getting closer to reality

Abstract

Next generation sequencing (NGS) denotes novel sequencing technologies that enable the generation of a large number of clonal sequences in a single sequencing run. NGS was initially introduced for whole genome sequencing and for quantitation of viral variants or genetic mutations in tumor tissues; more recently, the potential for high resolution HLA typing and high throughput analyses has been explored. It became clear that the complexity of the HLA system implicates new challenges, especially for bioinformatics. From an economical point of view, NGS is becoming increasingly attractive for HLA typing laboratories currently relying on Sanger based sequencing. Realizing the full potential of NGS will require the development of specifically adapted typing strategies and software algorithms. In the present review, three laboratories that were among the first to perform HLA-typing using different NGS platforms, the Roche 454, the Illumina Miseq and the Ion Torrent system, respectively, give an overview of these applications and point out advantages and limitations.

Erythrogene: a database for in-depth analysis of the extensive variation in 36 blood group systems in the 1000 Genomes Project

“…Blood group prediction based on NGS has been on the horizon for several years now, but interest was further sparked by introduction of this technology into tissue typing laboratories. 18 Earlier this year, NGS-based blood group and platelet phenotype predictions were reported for a single individual in a proof-of-principle study. 20 All blood group systems except XG were assessed, and .200 antigens were predicted, 17 of which were serologically confirmed.…”

“…16,17 Next-generation sequencing (NGS) has the potential to redress these limitations and is being implemented for HLA analysis in tissue typing reference laboratories. 18 Similarly, NGS is rapidly moving toward routine practice for patient and donor blood group typing, 19,20 yet a large worldwide survey of genomic blood group variation is lacking.…”

HLA typing: Conventional techniques v. next-generation sequencing

“…This study therefore indicates that the value of NGS over conventional techniques will only become significant in the context of HSCT and cord blood banking in SA when the number of samples increases to the point where NGS becomes more cost-effective than conventional techniques. [10] …”

Very high resolution single pass HLA genotyping using amplicon sequencing on the 454 next generation DNA sequencers: Comparison with Sanger sequencing

“…Together with software enhancements, it has the potential 42 to provide even higher resolution HLA typing. In recent years, various next generation sequencing platforms, 51 based on massively parallel clonal sequencing, have been used to 52 develop high resolution and high throughput HLA typing systems 53 [1][2][3][4][5][6][7][8][9][10][11][12]. In general, two basic strategies have been used: (1) an 54 amplicon sequencing approach focused on the highly polymorphic 55 regions (primarily exons) of HLA class I and class II genes [1][2][3][4][5][6], or 56 (2) long-range PCR of full or partial length individual HLA gene loci, 57 followed by fragmentation, shot-gun sequencing and assembly [7-58 11].…”