TY - JOUR
T1 - Development of a novel, genome subtraction-derived, sars-cov-2-specific covid-19-nsp2 real-time rt-pcr assay and its evaluation using clinical specimens
AU - Yip, Cyril Chik Yan
AU - Ho, Chi Chun
AU - Chan, Jasper Fuk Woo
AU - To, Kelvin Kai Wang
AU - Chan, Helen Shuk Ying
AU - Wong, Sally Cheuk Ying
AU - Leung, Kit Hang
AU - Fung, Agnes Yim Fong
AU - Ng, Anthony Chin Ki
AU - Zou, Zijiao
AU - Tam, Anthony Raymond
AU - Chung, Tom Wai Hin
AU - Chan, Kwok Hung
AU - Hung, Ivan Fan Ngai
AU - Cheng, Vincent Chi Chung
AU - Tsang, Owen Tak Yin
AU - Tsui, Stephen Kwok Wing
AU - Yuen, Kwok Yung
N1 - Publisher Copyright:
© 2020 by the authors. Licensee MDPI, Basel, Switzerland.
PY - 2020/4/1
Y1 - 2020/4/1
N2 - The pandemic novel coronavirus infection, Coronavirus Disease 2019 (COVID-19), has affected at least 190 countries or territories, with 465,915 confirmed cases and 21,031 deaths. In a containment-based strategy, rapid, sensitive and specific testing is important in epidemiological control and clinical management. Using 96 SARS-CoV-2 and 104 non-SARS-CoV-2 coronavirus genomes and our in-house program, GolayMetaMiner, four specific regions longer than 50 nucleotides in the SARS-CoV-2 genome were identified. Primers were designed to target the longest and previously untargeted nsp2 region and optimized as a probe-free real-time reverse transcription-polymerase chain reaction (RT-PCR) assay. The new COVID-19-nsp2 assay had a limit of detection (LOD) of 1.8 TCID50 /mL and did not amplify other human-pathogenic coronaviruses and respiratory viruses. Assay reproducibility in terms of cycle threshold (Cp) values was satisfactory, with the total imprecision (% CV) values well below 5%. Evaluation of the new assay using 59 clinical specimens from 14 confirmed cases showed 100% concordance with our previously developed COVID-19-RdRp/Hel reference assay. A rapid, sensitive, SARS-CoV-2-specific real-time RT-PCR assay, COVID-19-nsp2, was developed.
AB - The pandemic novel coronavirus infection, Coronavirus Disease 2019 (COVID-19), has affected at least 190 countries or territories, with 465,915 confirmed cases and 21,031 deaths. In a containment-based strategy, rapid, sensitive and specific testing is important in epidemiological control and clinical management. Using 96 SARS-CoV-2 and 104 non-SARS-CoV-2 coronavirus genomes and our in-house program, GolayMetaMiner, four specific regions longer than 50 nucleotides in the SARS-CoV-2 genome were identified. Primers were designed to target the longest and previously untargeted nsp2 region and optimized as a probe-free real-time reverse transcription-polymerase chain reaction (RT-PCR) assay. The new COVID-19-nsp2 assay had a limit of detection (LOD) of 1.8 TCID50 /mL and did not amplify other human-pathogenic coronaviruses and respiratory viruses. Assay reproducibility in terms of cycle threshold (Cp) values was satisfactory, with the total imprecision (% CV) values well below 5%. Evaluation of the new assay using 59 clinical specimens from 14 confirmed cases showed 100% concordance with our previously developed COVID-19-RdRp/Hel reference assay. A rapid, sensitive, SARS-CoV-2-specific real-time RT-PCR assay, COVID-19-nsp2, was developed.
KW - Clinical evaluation
KW - COVID-19
KW - COVID-19-nsp2 assay
KW - Genome subtraction
KW - GolayMetaMiner
KW - Nsp2
KW - Real-time RT-PCR
KW - SARS-CoV-2
KW - Sensitivity
KW - Specificity
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U2 - 10.3390/ijms21072574
DO - 10.3390/ijms21072574
M3 - Article
C2 - 32276333
AN - SCOPUS:85083206760
SN - 1661-6596
VL - 21
JO - International Journal of Molecular Sciences
JF - International Journal of Molecular Sciences
IS - 7
M1 - 2574
ER -