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

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

One goal among 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 we previously established as field standards. To enable simultaneous SARS-CoV-2 and microbial community profiling, we compare the relative performance of two total nucleic acid extraction protocols and our previously benchmarked protocol. We included a diverse panel of environmental and host-associated sample types, including body sites commonly swabbed for COVID-19 testing. Here we 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.