The Great Genotyper: A Graph-Based Method for Population Genotyping of Small and Structural Variants

1 Abstract Long-read sequencing (LRS) enables variant calling of high-quality structural variants (SVs). Genotypers of SVs utilize these precise call sets to increase the recall and precision of genotyping in short-read sequencing (SRS) samples. With the extensive growth in availabilty of SRS datasets in recent years, we should be able to calculate accurate population allele frequencies of SV. However, reprocessing hundreds of terabytes of raw SRS data to genotype new variants is impractical for population-scale studies, a computational challenge known as the N+1 problem. Solving this computational bottleneck is necessary to analyze new SVs from the growing number of pangenomes in many species, public genomic databases, and pathogenic variant discovery studies. To address the N+1 problem, we propose The Great Genotyper, a population genotyping workflow. Applied to a human dataset, the workflow begins by preprocessing 4.2K short-read samples of a total of 183TB raw data to create an 867GB Counting Colored De Bruijn Graph (CCDG). The Great Genotyper uses this CCDG to genotype a list of phased or unphased variants, leveraging the CCDG population information to increase both precision and recall. The Great Genotyper offers the same accuracy as the state-of-the-art genotypers with the addition of unprecedented performance. It took 100 hours to genotype 4.5M variants in the 4.2K samples using one server with 32 cores and 145GB of memory. A similar task would take months or even years using single-sample genotypers. The Great Genotyper opens the door to new ways to study SVs. We demonstrate its application in finding pathogenic variants by calculating accurate allele frequency for novel SVs. Also, a premade index is used to create a 4K reference panel by genotyping variants from the Human Pangenome Reference Consortium (HPRC). The new reference panel allows for SV imputation from genotyping microarrays. Moreover, we genotype the GWAS catalog and merge its variants with the 4K reference panel. We show 6.2K events of high linkage between the HPRC’s SVs and nearby GWAS SNPs, which can help in interpreting the effect of these SVs on gene functions. This analysis uncovers the detailed haplotype structure of the human fibrinogen locus and revives the pathogenic association of a 28 bp insertion in the FGA gene with thromboembolic disorders. Competing Interest Statement The authors have declared no competing interest. Funding Statement This study did not receive any funding Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: All source data were publicly available before the initiation of the study. ALl human sequencing samples can be freely obtained from: https://www.internationalgenome.org/data-portal/sample The human pangenome is available at: https://zenodo.org/records/6797328 The Clinvar variants are available at: https://zenodo.org/records/6797328 The Genome in a bottle datasets are available at: https://ftp.ncbi.nlm.nih.gov/ReferenceSamples/giab/release/AshkenazimTrio/HG002_NA24385_son/NISTv4.2.1/GRCh38/ https://ftp.ncbi.nlm.nih.gov/ReferenceSamples/giab/release/AshkenazimTrio/HG002_NA24385_son/NISTv4.2.1/GRCh38/ The human Gwas catalog: https://www.ebi.ac.uk/gwas/downloads I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes Data Availability All data produced in the present work are contained in the manuscript