Open AccessDetection of SARS-CoV-2 RNA by multiplex RT-qPCR
Author(s) -
Eriko Kudo,
Benjamin Israelow,
Chantal B.F. Vogels,
Peiwen Lu,
Anne L. Wyllie,
Maria Tokuyama,
Arvind Venkataraman,
Doug E. Brackney,
Isabel M. Ott,
Mary E. Petrone,
Rebecca Earnest,
Sarah Lapidus,
M. Catherine Muenker,
Adam J. Moore,
Arnau CasanovasMassana,
Saad B. Omer,
Charles S. Dela Cruz,
Shelli Farhadian,
Albert I. Ko,
Nathan D. Grubaugh,
Akiko Iwasaki
Publication year - 2020
Publication title -
plos biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 4.127
H-Index - 271
eISSN - 1545-7885
pISSN - 1544-9173
DOI - 10.1371/journal.pbio.3000867
Subject(s) - multiplex , biology , virology , real time polymerase chain reaction , covid-19 , molecular diagnostics , reverse transcription polymerase chain reaction , multiplex polymerase chain reaction , primer (cosmetics) , coronavirus , microbiology and biotechnology , computational biology , polymerase chain reaction , gene , gene expression , bioinformatics , genetics , infectious disease (medical specialty) , disease , medicine , pathology , chemistry , organic chemistry
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.