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Anal Biochem ; 641: 114565, 2022 03 15.
Article in English | MEDLINE | ID: covidwho-1632512

ABSTRACT

Polymerase chain reaction (PCR) is the most widely used method for nucleic acids amplification. To date, a huge number of versatile PCR techniques have been developed. One of the relevant goals is to shorten PCR duration, which can be achieved in several ways. Here, we report on the results regarding nucleic acids amplification by convective PCR (cPCR) in standard 0.2 ml polypropylene microtubes. The following conditions were found to be optimal for such amplification: 1) 70 µl reaction volume, 2) the supply of external temperature 145°Ð¡ for the denaturation zone and 0°Ð¡ for the annealing zone, 3) ∼30° inclination of the microtube main axis, 4) the use of nearby primers, and 5) duration of the reaction 15-20 min. At these conditions, the amplification products are accumulated in an amount sufficient to be registered by gel electrophoresis, and high sensitivity of the reaction comparable to that of conventional PCR is achieved. cPCR provided the reliable detection of SARS-CoV-2 coronavirus RNA isolated from nasopharyngeal swabs of COVID-19 patients.


Subject(s)
COVID-19 Nucleic Acid Testing/instrumentation , COVID-19/diagnosis , Polymerase Chain Reaction/instrumentation , SARS-CoV-2/isolation & purification , COVID-19/virology , COVID-19 Nucleic Acid Testing/economics , COVID-19 Nucleic Acid Testing/methods , Convection , Humans , Polymerase Chain Reaction/economics , Polymerase Chain Reaction/methods , RNA, Viral/analysis , RNA, Viral/genetics , SARS-CoV-2/genetics , Temperature , Time Factors
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