A comparison of DNA/RNA extraction protocols for high-throughput sequencing of microbial communities

Justin P Shaffer; Clarisse Marotz; Pedro Belda-Ferre; Cameron Martino; Stephen Wandro; Mehrbod Estaki; Rodolfo A Salido; Carolina S Carpenter; Livia S Zaramela; Jeremiah J Minich; MacKenzie Bryant; Karenina Sanders; Serena Fraraccio; Gail Ackermann; Gregory Humphrey; Austin D Swafford; Sandrine Miller-Montgomery; Rob Knight

doi:10.2144/btn-2020-0153

BioTechniques (Mar 2021)

A comparison of DNA/RNA extraction protocols for high-throughput sequencing of microbial communities

Justin P Shaffer,
Clarisse Marotz,
Pedro Belda-Ferre,
Cameron Martino,
Stephen Wandro,
Mehrbod Estaki,
Rodolfo A Salido,
Carolina S Carpenter,
Livia S Zaramela,
Jeremiah J Minich,
MacKenzie Bryant,
Karenina Sanders,
Serena Fraraccio,
Gail Ackermann,
Gregory Humphrey,
Austin D Swafford,
Sandrine Miller-Montgomery,
Rob Knight

Affiliations

Justin P Shaffer: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Clarisse Marotz: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Pedro Belda-Ferre: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Cameron Martino: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Stephen Wandro: 2Center for Microbiome Innovation, University of California, San Diego, La Jolla, CA, USA
Mehrbod Estaki: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Rodolfo A Salido: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Carolina S Carpenter: 2Center for Microbiome Innovation, University of California, San Diego, La Jolla, CA, USA
Livia S Zaramela: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Jeremiah J Minich: 4Marine Biology Research Division, University of California, San Diego, La Jolla, CA, USA
MacKenzie Bryant: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Karenina Sanders: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Serena Fraraccio: 2Center for Microbiome Innovation, University of California, San Diego, La Jolla, CA, USA
Gail Ackermann: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Gregory Humphrey: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA
Austin D Swafford: 2Center for Microbiome Innovation, University of California, San Diego, La Jolla, CA, USA
Sandrine Miller-Montgomery: 2Center for Microbiome Innovation, University of California, San Diego, La Jolla, CA, USA
Rob Knight: 1Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA

DOI: https://doi.org/10.2144/btn-2020-0153
Journal volume & issue: Vol. 70, no. 3
pp. 149 – 159

Abstract

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One goal of microbial ecology researchers is to capture the maximum amount of information from all organisms in a sample. The recent COVID-19 pandemic, caused by the RNA virus SARS-CoV-2, has highlighted a gap in traditional DNA-based protocols, including the high-throughput methods the authors previously established as field standards. To enable simultaneous SARS-CoV-2 and microbial community profiling, the authors compared the relative performance of two total nucleic acid extraction protocols with the authors' previously benchmarked protocol. The authors included a diverse panel of environmental and host-associated sample types, including body sites commonly swabbed for COVID-19 testing. Here the authors present results comparing the cost, processing time, DNA and RNA yield, microbial community composition, limit of detection and well-to-well contamination between these protocols.

Published in BioTechniques

ISSN: 0736-6205 (Print); 1940-9818 (Online)
Publisher: Taylor & Francis Group
Country of publisher: United Kingdom
LCC subjects: Science: Biology (General)
Website: https://www.tandfonline.com/journals/ibtn20

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