Resource-efficient internally controlled in-house real-time PCR detection of SARS-CoV-2
The reliable detection of SARS-CoV-2 has become one of the most important contributions to COVID-19 crisis management. With the publication of the first sequences of SARS-CoV-2, several diagnostic PCR assays have been developed and published. In addition to in-house assays the market was flooded wit...
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Veröffentlicht in: | Virology journal 2021-06, Vol.18 (1), p.110-11, Article 110 |
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Sprache: | eng |
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Zusammenfassung: | The reliable detection of SARS-CoV-2 has become one of the most important contributions to COVID-19 crisis management. With the publication of the first sequences of SARS-CoV-2, several diagnostic PCR assays have been developed and published. In addition to in-house assays the market was flooded with numerous commercially available ready-to-use PCR kits, with both approaches showing alarming shortages in reagent supply.
Here we present a resource-efficient in-house protocol for the PCR detection of SARS-CoV-2 RNA in patient specimens (RKI/ZBS1 SARS-CoV-2 protocol).
Two duplex one-step real-time RT-PCR assays are run simultaneously and provide information on two different SARS-CoV-2 genomic regions. Each one is duplexed with a control that either indicates potential PCR inhibition or proves the successful extraction of nucleic acid from the clinical specimen.
Limit of RNA detection for both SARS-CoV-2 assays is below 10 genomes per reaction. The protocol enables testing specimens in duplicate across the two different SARS-CoV-2 PCR assays, saving reagents by increasing testing capacity. The protocol can be run on various PCR cyclers with several PCR master mix kits.
The presented RKI/ZBS1 SARS-CoV-2 protocol represents a cost-effective alternative in times of shortages when commercially available ready-to-use kits may not be available or affordable. |
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ISSN: | 1743-422X 1743-422X |
DOI: | 10.1186/s12985-021-01559-3 |