PROJECT SUMMARY The University of Louisville’s Sequencing Technology Center (STC) is requesting support to acquire a Pacific Biosciences Sequel IIe sequencing system. The requested system would be the second Sequel IIe system in the Center, as demand from the diverse STC user base, comprised of regional, national, and international extramurally funded investigators has far exceeded the capacity of our initial, single instrument. Since launching in 2021, the STC has assisted in the planning, preparation, and execution of many projects leveraging the power of long read, single molecule, real-time (SMRT) sequencing for projects that use both standard high fidelity (HiFi) workflows, and those that involve development of custom, innovative targeted methods to elucidate complex genomic loci or communities with phased, highly accurate long reads. Unlike other sequencing systems on the market, the Sequel IIe system has the combination of long read capabilities (150 bp – 100 kb) and the highest accuracy available from any NGS- and single molecule-sequencing instrument (ranging from Q30, 99.9% - to Q60, 99.9999%, across our projects), both of which are critical for meeting the needs of our Major, Minor, and service users. The STC is well established and already has the infrastructure and operational capabilities to implement this requested second Sequel IIe system, which is absolutely necessary for meeting the needs of our growing user base, Major User research projects, and further genomics methods development using the power of SMRT sequencing technology. The University of Louisville, particularly the Office of Research and Innovation, is dedicated to supporting the requested system through financial and infrastructure investment into the STC. From the generation of new reference genomes to targeted characterization of viral reservoirs and immune loci, and critically important SARS-CoV-2 pathogen surveillance across the Commonwealth of Kentucky, the STC facility has and will use its expertise with SMRT sequencing and the proposed new Sequel IIe system to work closely with local and national experts to facilitate cutting-edge genomics projects across a wide variety of research areas with the major goal of promoting and inspiring research and innovation in support of the growth of the research enterprise at the University of Louisville.

PROJECT NARRATIVE High fidelity (HiFi) reads generated with single molecule, real time (SMRT) sequencing are the foundational data type needed to generate highly accurate, contiguous whole genome de novo assemblies, and are incredibly powerful when used in a targeted fashion to unambiguously resolve haplotype-phased complex genomic loci. As nucleic acid sequencing technologies continue to develop and improve, we have the responsibility and need to provide investigators with the state-of-the-art tools required to perform cutting-edge genomics research in a timely and cost-effective manner. The goal of this proposal is to obtain support for the purchase of a second Pacific Biosciences Sequel IIe sequencing system for our Sequencing Technology Center, which will provide much needed additional bandwidth and throughput for funded NIH projects, significantly enhancing the biomedical research programs at University of Louisville and our regional and national collaborative partners.