Статья

Detection of SARS-CoV-2 RNA by a Multiplex Reverse-Transcription Loop-Mediated Isothermal Amplification Coupled with Melting Curves Analysis.

I. Oscorbin, G. Shevelev, K. Pronyaeva, A. Stepanov, D. Shamovskaya, O. Mishukova, D. Pyshnyi, M. Filipenko,
2021

Loop-mediated isothermal amplification (LAMP) is a method of nucleic acid amplification that is more stable and resistant to DNA amplification inhibitors than conventional PCR. LAMP multiplexing with reverse transcription allows for the single-tube amplification of several RNA fragments, including an internal control sample, which provides the option of controlling all analytical steps. We developed a method of SARS-CoV-2 viral RNA detection based on multiplex reverse-transcription LAMP, with single-tube qualitative analysis of SARS-CoV-2 RNA and MS2 phage used as a control RNA. The multiplexing is based on the differences in characteristic melting peaks generated during the amplification process. The developed technique detects at least 20 copies of SARS-CoV-2 RNA per reaction on a background of 12,000 MS2 RNA copies. The total time of analysis does not exceed 40 min. The method validation, performed on 125 clinical samples of patients' nasal swabs, showed a 97.6% concordance rate with the results of real-time (RT)-PCR assays. The developed multiplexed LAMP can be employed as an alternative to PCR in diagnostic practice to save personnel and equipment time.

Цитирование

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Версии

  • 1. Version of Record от 2021-05-27

Метаданные

Об авторах
  • I. Oscorbin
    Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences (ICBFM SB RAS), 8 Lavrentiev Avenue, 630090 Novosibirsk, Russia.
  • G. Shevelev
    Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences (ICBFM SB RAS), 8 Lavrentiev Avenue, 630090 Novosibirsk, Russia.
  • K. Pronyaeva
    Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences (ICBFM SB RAS), 8 Lavrentiev Avenue, 630090 Novosibirsk, Russia.
  • A. Stepanov
    Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences (ICBFM SB RAS), 8 Lavrentiev Avenue, 630090 Novosibirsk, Russia.
  • D. Shamovskaya
    Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences (ICBFM SB RAS), 8 Lavrentiev Avenue, 630090 Novosibirsk, Russia.
  • O. Mishukova
    Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences (ICBFM SB RAS), 8 Lavrentiev Avenue, 630090 Novosibirsk, Russia.
  • D. Pyshnyi
    Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences (ICBFM SB RAS), 8 Lavrentiev Avenue, 630090 Novosibirsk, Russia.
  • M. Filipenko
    Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences (ICBFM SB RAS), 8 Lavrentiev Avenue, 630090 Novosibirsk, Russia.
Название журнала
  • International Journal of Molecular Sciences
Том
  • 22
Выпуск
  • 11
Страницы
  • 5743
Издатель
  • MDPI
Тип документа
  • journal article
Тип лицензии Creative Commons
  • CC BY
Правовой статус документа
  • Свободная лицензия
Источник
  • dimensions