Homo sapiens Raw sequence reads

Human Leukocyte Antigen (HLA) genes are pivotal in numerous immunological applications. Given their immense polymorphism, achieving accurate high-throughput HLA typing is challenging. This study endeavors to harness the Human Pangenome Reference Consortium (HPRC) resources as a potential benchmark for HLA reference materials. We meticulously annotated specific four-field resolution alleles for 11 HLA genes (including HLA-A, -B, -C, -DPA1, -DPB1, -DQA1, -DQB1, -DRB1, -DRB3, -DRB4 and -DRB5) from 44 high-quality HPRC personal genome assemblies. For sequencing, we crafted HLA-specific probes, conducting capture-based targeted sequencing on the HPRC cohort's genomic DNA, ensuring focused and comprehensive coverage of the HLA region of interest. Concurrently, we utilized publicly available short-read whole genome sequencing (WGS) data from identical samples, offering a comparative perspective. To decipher this vast sequencing data, we employed six distinct software tools: HLA-VBseq, HISAT-genotype, SpecHLA, T1K, QzType, and DRAGEN. Each tool offered unique capabilities and algorithms for HLA genotyping, allowing for comprehensive analysis and validation of results. We then compared these outcomes against benchmarks derived from the personal genome assemblies. Our findings present a comprehensive, four-field resolution HLA allele annotation for the 44 HPRC samples. Significantly, our innovative targeted next-generation sequencing (NGS) approach for HLA genes showed superior accuracy over conventional short-read WGS. An integrated analysis involving QzType, T1K, and DRAGEN outshined, hitting a flawless 100% accuracy for all 11 HLA genes. In conclusion, our study underscores the combination of targeted short-read sequencing and astute computational analysis as a robust approach for HLA genotyping. Furthermore, the HPRC cohort emerges as a prime gold-standard reference in this realm.