METHODS article
Front. Genet.
Sec. Genomic Assay Technology
Volume 16 - 2025 | doi: 10.3389/fgene.2025.1587691
This article is part of the Research TopicAdvancements in Sequencing Technologies for Epigenomic and Transcriptomic Analysis: From Bulk to Single-Cell ResolutionView all 3 articles
High-Throughput PacBio Library Preparation and Sequencing Techniques for Genomic DNA and TNA
Provisionally accepted
- Centers for Disease Control and Prevention (Georgia), Atlanta, Georgia
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PacBio Sequencing is an effective tool for achieving long read lengths to close gaps in whole genome assemblies. Limited information is available for automated, highthroughput library preparation workflows for PacBio sequencing. This paper describes two high throughput methods, one for producing barcoded genomic DNA libraries and one for barcoded cDNA and subsequent library preparation, as well as pooling and sequencing on the Sequel II instrument. Test sequencing was performed on 380 genomic DNA samples, of which 28 (7.4%) failed sequencing and needed to be repeated. Test sequencing was performed on 384 Threose nucleic acid samples of which 0 failed sequencing and 49 (12.8%) had a genome coverage <90%. These methods were used to generate and report results for ~2,000 genomic DNA and ~30,000 cDNA genome sequences from clinical specimens in 2023.
Keywords: WGS - whole-genome sequencing, high throughput, PacBio, Automation, methods, gDNA, COVID, Sequencing
Received: 26 Mar 2025; Accepted: 18 Jul 2025.
Copyright: © 2025 Bentz, Clines, Padilla, Horn, Lee and Sheth. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Meghan L Bentz, Centers for Disease Control and Prevention (Georgia), Atlanta, Georgia
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