In this CSHL Biology of Genomes 2021 virtual workshop, Aaron Wenger from PacBio discusses examples of how advances in highly accurate long-read (HiFi) sequencing have enabled exciting developments in human genome research, including sequencing the genomes of 100 individuals with unexplained diseases.
In this talk, Dr. Wenger describes how whole-genome sequencing (WGS) with accurate, long HiFi reads identify all the variations found with short reads plus small variants in difficult-to-map regions and structural variants across the genome. He further explains how HiFi reads also support direct phasing of variants into haplotypes. Researchers worldwide apply HiFi reads to explain rare disease cases unsolved by other technologies. Improvements in workflow, reliability, cost and throughput support the routine application of HiFi reads in large studies today and open a future of HiFi genomes as a standard tool for rare disease researchers.
Short-read genome-wide sequencing for molecular diagnosis has revolutionized pediatric rare disease care in the past decade. However, most families remain without specific knowledge of the cause of their child’s illness. We seek to understand how long-read sequencing (HiFi sequencing) and functional genomics can fill the gaps and identify most causes of genetic disease. Dr. Pastinen describes a health-system-wide initiative to translate the latest research approaches to end the diagnostic “odyssey” affecting rare disease families, observing an expanded range of variation and enhanced interpretation of known variation by integrating HiFi data to unsolved rare disease cases.
Over the past few years, many tools have been developed to enable comprehensive variant detection from PacBio HiFi reads. This talk describes a flexible, modular workflow for variant detection and prioritization from HiFi whole-genome sequencing data, including open-source tools for quality control, alignment, small variant detection, and phasing, structural variant detection, genotyping of tandem repeats, and de novo assembly. This pipeline is available on GitHub as a Snakemake workflow and has been adapted into a Cromwell WDL workflow by Microsoft Genomics.
Alexander Hoischen’s research group ‘Genomic Technologies and Immuno-Genomics’ has expertise in the identification of rare disease genes using the latest genomics tools, with a recent particular focus on immune-related disease genes. His group has been the first to identify a disease causing dominant de novo mutation for a Mendelian disorder by exome sequencing, followed by the identification of several disease genes for rare diseases. Following a six-month research stint in 2013 in the laboratories of collaborators, Prof. Eichler and Prof. Shendure (UW, Seattle; USA), Dr. Hoischen established the latest technology for accurate and large-scale targeted re-sequencing (smMIPs) in Nijmegen. Recently,…
In this talk, Christine Lambert describes a simple and scalable workflow for generating high-quality HiFi reads appropriate for comprehensive variant detection (SNVs, Indels, SVs) and de novo assembly. Using this workflow, multiple samples can be processed manually (up to 16) with a multichannel pipet and strip tubes. Up to 96 samples can be processed on an automated liquid handler such as the Sciclone Liquid Handler Workstation. She also describes solutions for high-throughput SMRTbell library construction for generating HiFi reads on the Sequel IIe System.
In this talk, Dr. Ekholm describes how long-read sequencing is being incorporated for rare Mendelian disease research, why high accuracy matters in long-read sequencing, what can be detected with HiFi reads that is missed with standard sequencing methods, and finally, how long-read sequencing can help increase the solve rates for rare and Mendelian diseases.
In this talk, Dr. Matsumoto describes his research of a family with syndromic intellectual disability. Trio-base exome analysis could not find any culprit mutation. Therefore, he and his team applied trio-based HiFi long-read WGS using two flowcells for a patient and one flowcell each for her father and mother. Through systematic variant filtering, they could find a 12-kb copy neutral inversion disrupting a causative gene. In addition, they could confirm that the de novo inversion occurred on the paternal chromosome through the haplotype phasing. These data demonstrate the utility of HiFi long-read WGS in solving patients with rare diseases.