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?:abstract
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The SARS-CoV-2 emerged in December 2019 and quickly spread around the world forcing global health authorities to develop protocols for its diagnosis. Here we report dimer formation in the N2 primers-probe set (CDC 2019-nCoV Real-Time RT-PCR) used in diagnostic routine, and propose alternatives to reduce dimerization events. Late unspecific amplifications were visualized in 56.4% and 57.1% of negative samples and no-template control, respectively, but not in positive samples and positive control. In silico analysis and gel electrophoresis confirm the dimer formation. The RT-qPCR parameters were optimized and the late unspecific amplifications decreased to 11.5% in negative samples and no-template control. The adjustment of PCR parameters was essential to reduce the risk of false positives results and to avoid inclusive results that require repetition of tests, which increases the costs and generates delays in results or even unnecessary requests for new samples.
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?:doi
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10.1016/j.ijid.2020.10.079
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document_parses/pdf_json/a1a6cc1e8ada302dcf269e1e4ef8751046ee3e0e.json
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?:title
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Adjusting RT-qPCR conditions to avoid unspecific amplification in SARS-CoV-2 diagnostic
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