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Olivar: towards automated variant aware primer design for multiplex tiled amplicon sequencing of pathogens

Michael X. Wang, Esther G. Lou, Nicolae Sapoval, Eddie Kim, Prashant Kalvapalle, Bryce Kille, R. A. Leo Elworth, Yunxi Liu, Yilei Fu, Lauren B. Stadler () and Todd J. Treangen ()
Additional contact information
Michael X. Wang: Rice University
Esther G. Lou: Rice University
Nicolae Sapoval: Rice University
Eddie Kim: Rice University
Prashant Kalvapalle: Rice University
Bryce Kille: Rice University
R. A. Leo Elworth: Rice University
Yunxi Liu: Rice University
Yilei Fu: Rice University
Lauren B. Stadler: Rice University
Todd J. Treangen: Rice University

Nature Communications, 2024, vol. 15, issue 1, 1-12

Abstract: Abstract Tiled amplicon sequencing has served as an essential tool for tracking the spread and evolution of pathogens. Over 15 million complete SARS-CoV-2 genomes are now publicly available, most sequenced and assembled via tiled amplicon sequencing. While computational tools for tiled amplicon design exist, they require downstream manual optimization both computationally and experimentally, which is slow and costly. Here we present Olivar, a first step towards a fully automated, variant-aware design of tiled amplicons for pathogen genomes. Olivar converts each nucleotide of the target genome into a numeric risk score, capturing undesired sequence features that should be avoided. In a direct comparison with PrimalScheme, we show that Olivar has fewer mismatches overlapping with primers and predicted PCR byproducts. We also compare Olivar head-to-head with ARTIC v4.1, the most widely used primer set for SARS-CoV-2 sequencing, and show Olivar yields similar read mapping rates (~90%) and better coverage to the manually designed ARTIC v4.1 amplicons. We also evaluate Olivar on real wastewater samples and found that Olivar has up to 3-fold higher mapping rates while retaining similar coverage. In summary, Olivar automates and accelerates the generation of tiled amplicons, even in situations of high mutation frequency and/or density. Olivar is available online as a web application at https://olivar.rice.edu and can be installed locally as a command line tool with Bioconda. Source code, installation guide, and usage are available at https://github.com/treangenlab/Olivar .

Date: 2024
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DOI: 10.1038/s41467-024-49957-9

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