Whole-genome variant detection in long-read sequencing data from ultra-low input tumor samples

Long-read sequencing (LRS) provides a more complete view of the genome than short-read sequencing, with advantages in haplotype phasing, structural variant detection, and access to difficult-to-map regions. One limitation of LRS has been high input DNA requirements, with several micrograms required per sample. Here we evaluate two methods of amplification-based long-read, whole-genome sequencing: Ultra-Low Input HiFi (ULI-HiFi) sequencing and droplet multiple displacement amplification (dMDA) sequencing. When these methods were benchmarked against a reference sample, HG002, we observed high precision and recall of single nucleotide variants (SNVs) with ULI-HiFi compared to the dMDA-amplified samples. ULI-HiFi successfully illuminated several medically-important genes that were poorly covered with short-read DNA sequencing. Because ULI-HiFi only requires ~10 nanograms of DNA, we extended ULI-HiFi to sequence adenocarcinoma and polyp samples from a patient with familial adenomatous polyposis, a hereditary form of colorectal cancer. This analysis revealed SNVs and structural variants not detected by short-read sequencing technologies. ULI-HiFi will enable the improved characterization of genetic variants in dark regions of the genome from precious clinical samples, which will allow for a better understanding of human disease and more efficacious precision medicines.