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Biochem Biophys Res Commun. 2003 Dec 26;312(4):1290-6.

A real-time PCR for SARS-coronavirus incorporating target gene pre-amplification.

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  • 1Key Laboratory of Neuroscience, Neuroscience Research Institute, Peking University and Department of Neurobiology, Peking University Health Science Center, 38 Xue Yuan Road, 100083, Beijing, China.

Abstract

An enhanced polymerase chain reaction (PCR) assay to detect the coronavirus associated with severe acute respiratory syndrome (SARS-CoV) was developed in which a target gene pre-amplification step preceded TaqMan real-time fluorescent PCR. Clinical samples were collected from 120 patients diagnosed as suspected or probable SARS cases and analyzed by conventional PCR followed by agarose gel electrophoresis, conventional TaqMan real-time PCR, and our enhanced TaqMan real-time PCR assays. An amplicon of the size expected from SARS-CoV was obtained from 28/120 samples using the enhanced real-time PCR method. Conventional PCR and real-time PCR alone identified fewer SARS-CoV positive cases. Results were confirmed by viral culture in 3/28 cases. The limit of detection of the enhanced real-time PCR method was 10(2)-fold higher than the standard real-time PCR assay and 10(7)-fold higher than conventional PCR methods. The increased sensitivity of the assay may help control the spread of the disease during future SARS outbreaks.

PMID:
14652014
[PubMed - indexed for MEDLINE]
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