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A Melting Curve-Based Multiplex RT-qPCR Assay for Simultaneous Detection of Four Human Coronaviruses

International journal of molecular sciences, 2016-11, Vol.17 (11), p.1880-1880 [Peer Reviewed Journal]

Copyright MDPI AG 2016 ;2016 by the authors; licensee MDPI, Basel, Switzerland. 2016 ;ISSN: 1422-0067 ;ISSN: 1661-6596 ;EISSN: 1422-0067 ;DOI: 10.3390/ijms17111880 ;PMID: 27886052

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Title:
A Melting Curve-Based Multiplex RT-qPCR Assay for Simultaneous Detection of Four Human Coronaviruses
Author: Wan, Zhenzhou ; Zhang, Ya'nan ; He, Zhixiang ; Liu, Jia ; Lan, Ke ; Hu, Yihong ; Zhang, Chiyu
Subjects: Acute Disease ; Coronavirus - genetics ; Coronavirus - isolation & purification ; Coronavirus 229E, Human - genetics ; Coronavirus 229E, Human - isolation & purification ; Coronavirus NL63, Human - genetics ; Coronavirus NL63, Human - isolation & purification ; Coronavirus OC43, Human - genetics ; Coronavirus OC43, Human - isolation & purification ; Diagnosis, Differential ; DNA Primers - chemical synthesis ; DNA Primers - metabolism ; DNA, Complementary - biosynthesis ; DNA, Complementary - genetics ; Fluorescent Dyes - chemistry ; Humans ; Limit of Detection ; Multiplex Polymerase Chain Reaction - methods ; Multiplex Polymerase Chain Reaction - standards ; Nucleic Acid Denaturation ; Organic Chemicals - chemistry ; Polymerase chain reaction ; Real-Time Polymerase Chain Reaction - methods ; Real-Time Polymerase Chain Reaction - standards ; Respiratory diseases ; Respiratory Tract Infections - diagnosis ; Respiratory Tract Infections - virology ; RNA, Viral - genetics ; RNA, Viral - isolation & purification ; Viral infections
Is Part Of: International journal of molecular sciences, 2016-11, Vol.17 (11), p.1880-1880
Description: Human coronaviruses HCoV-OC43, HCoV-229E, HCoV-NL63 and HCoV-HKU1 are common respiratory viruses associated with acute respiratory infection. They have a global distribution. Rapid and accurate diagnosis of HCoV infection is important for the management and treatment of hospitalized patients with HCoV infection. Here, we developed a melting curve-based multiplex RT-qPCR assay for simultaneous detection of the four HCoVs. In the assay, SYTO 9 was used to replace SYBR Green I as the fluorescent dye, and GC-modified primers were designed to improve the melting temperature (Tm) of the specific amplicon. The four HCoVs were clearly distinguished by characteristic melting peaks in melting curve analysis. The detection sensitivity of the assay was 3 × 10² copies for HCoV-OC43, and 3 × 10¹ copies for HCoV-NL63, HCoV-229E and HCoV-HKU1 per 30 μL reaction. Clinical evaluation and sequencing confirmation demonstrated that the assay was specific and reliable. The assay represents a sensitive and reliable method for diagnosis of HCoV infection in clinical samples.
Publisher: Switzerland: MDPI AG
Language: English
Identifier: ISSN: 1422-0067
ISSN: 1661-6596
EISSN: 1422-0067
DOI: 10.3390/ijms17111880
PMID: 27886052
Source: MEDLINE
PubMed Central
Coronavirus Research Database
ProQuest Central
DOAJ Directory of Open Access Journals

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A Melting Curve-Based Multiplex RT-qPCR Assay for Simultaneous Detection of Four Human Coronaviruses

Copyright MDPI AG 2016 ;2016 by the authors; licensee MDPI, Basel, Switzerland. 2016 ;ISSN: 1422-0067 ;ISSN: 1661-6596 ;EISSN: 1422-0067 ;DOI: 10.3390/ijms17111880 ;PMID: 27886052

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