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Detection of SARS-CoV-2 RNA by multiplex RT-qPCR.
Kudo, Eriko; Israelow, Benjamin; Vogels, Chantal B F; Lu, Peiwen; Wyllie, Anne L; Tokuyama, Maria; Venkataraman, Arvind; Brackney, Doug E; Ott, Isabel M; Petrone, Mary E; Earnest, Rebecca; Lapidus, Sarah; Muenker, M Catherine; Moore, Adam J; Casanovas-Massana, Arnau; Omer, Saad B; Dela Cruz, Charles S; Farhadian, Shelli F; Ko, Albert I; Grubaugh, Nathan D; Iwasaki, Akiko.
  • Kudo E; Department of Immunobiology, Yale School of Medicine, New Haven, Connecticut, United States of America.
  • Israelow B; Department of Immunobiology, Yale School of Medicine, New Haven, Connecticut, United States of America.
  • Vogels CBF; Department of Internal Medicine, Section of Infectious Diseases, Yale School of Medicine, New Haven, Connecticut, United States of America.
  • Lu P; Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, Connecticut, United States of America.
  • Wyllie AL; Department of Immunobiology, Yale School of Medicine, New Haven, Connecticut, United States of America.
  • Tokuyama M; Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, Connecticut, United States of America.
  • Venkataraman A; Department of Immunobiology, Yale School of Medicine, New Haven, Connecticut, United States of America.
  • Brackney DE; Department of Immunobiology, Yale School of Medicine, New Haven, Connecticut, United States of America.
  • Ott IM; The Connecticut Agricultural Experiment Station, Department of Environmental Sciences, New Haven, Connecticut, United States of America.
  • Petrone ME; Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, Connecticut, United States of America.
  • Earnest R; Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, Connecticut, United States of America.
  • Lapidus S; Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, Connecticut, United States of America.
  • Muenker MC; Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, Connecticut, United States of America.
  • Moore AJ; Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, Connecticut, United States of America.
  • Casanovas-Massana A; Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, Connecticut, United States of America.
  • Dela Cruz CS; Department of Internal Medicine, Section of Infectious Diseases, Yale School of Medicine, New Haven, Connecticut, United States of America.
  • Farhadian SF; Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, Connecticut, United States of America.
  • Ko AI; Yale Institute of Global Health, New Haven, Connecticut, United States of America.
  • Grubaugh ND; Yale School of Nursing, New Haven, Connecticut, United States of America.
  • Iwasaki A; Section of Pulmonary, Critical Care, and Sleep Medicine, Department of Internal Medicine, Yale School of Medicine, New Haven, Connecticut, United States of America.
PLoS Biol ; 18(10): e3000867, 2020 10.
Article in English | MEDLINE | ID: covidwho-901993
Preprint
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ABSTRACT
The current quantitative reverse transcription PCR (RT-qPCR) assay recommended for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) testing in the United States requires analysis of 3 genomic targets per sample 2 viral and 1 host. To simplify testing and reduce the volume of required reagents, we devised a multiplex RT-qPCR assay to detect SARS-CoV-2 in a single reaction. We used existing N1, N2, and RP primer and probe sets by the Centers for Disease Control and Prevention, but substituted fluorophores to allow multiplexing of the assay. The cycle threshold (Ct) values of our multiplex RT-qPCR were comparable to those obtained by the single assay adapted for research purposes. Low copy numbers (≥500 copies/reaction) of SARS-CoV-2 RNA were consistently detected by the multiplex RT-qPCR. Our novel multiplex RT-qPCR improves upon current single diagnostics by saving reagents, costs, time, and labor.
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Full text: Available Collection: International databases Database: MEDLINE Main subject: Pneumonia, Viral / Reagent Kits, Diagnostic / RNA, Viral / Coronavirus Infections / Clinical Laboratory Techniques / Reverse Transcriptase Polymerase Chain Reaction / Multiplex Polymerase Chain Reaction / Betacoronavirus Type of study: Diagnostic study / Observational study / Prognostic study Limits: Humans Country/Region as subject: North America Language: English Journal: PLoS Biol Journal subject: Biology Year: 2020 Document Type: Article Affiliation country: JOURNAL.PBIO.3000867

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Full text: Available Collection: International databases Database: MEDLINE Main subject: Pneumonia, Viral / Reagent Kits, Diagnostic / RNA, Viral / Coronavirus Infections / Clinical Laboratory Techniques / Reverse Transcriptase Polymerase Chain Reaction / Multiplex Polymerase Chain Reaction / Betacoronavirus Type of study: Diagnostic study / Observational study / Prognostic study Limits: Humans Country/Region as subject: North America Language: English Journal: PLoS Biol Journal subject: Biology Year: 2020 Document Type: Article Affiliation country: JOURNAL.PBIO.3000867