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In Vitro System for the Detection of Prostate Cancer Markers via Loop-Mediated Isothermal Amplification

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Abstract

Loop-mediated isothermal amplification (LAMP) of nucleic acids enables the detection of amplification products 10–20 min after the beginning of the reaction. An intraoperative method for the detection of metastases in lymph nodes based on LAMP, one-step nucleic acid amplification (OSNA), allows the rapid detection of the epithelial-specific marker gene KRT19 in lymph nodes. The standard protocol for OSNA was developed more than 10 years ago to detect breast cancer metastases in lymph nodes. Since then, a new version of the key enzyme involved in the reaction (Bst-polymerase) was obtained, but its use in OSNA has remained unexplored. Moreover, the time has come to apply OSNA to the detection of other cancer types, in particular, prostate cancer. The first step is to create an in vitro system that allows LAMP to be carried out on prostate-cancer cell lines. In this work, the LAMP protocol was developed for the new Bst 3.0 polymerase with optimized dNTP concentrations and without reverse transcriptase, and cell lines were selected (DU-145 for prostate cancer and Molt-4 for negative control). As a result, a new, in vitro system for the LAMP-based detection of prostate cancer with the marker gene KRT19 was developed.

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Funding

This work was supported by a grant from the President of the Russian Federation for State Support of Young Russian Scientists (MK-1906.2019.4).

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Authors and Affiliations

  1. Hertsen Moscow Oncology Research Institute, National Center of Medical Radiological Research, 125284, Moscow, Russia

    K. A. Fomicheva & J. A. Makarova

  2. Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, 119991, Moscow, Russia

    J. A. Makarova

  3. Faculty of Biology and Biotechnologies, Higher School of Economics National Research University, 101000, Moscow, Russia

    J. A. Makarova

Authors
  1. K. A. Fomicheva
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  2. J. A. Makarova
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Correspondence to K. A. Fomicheva or J. A. Makarova.

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The authors declare that they have no conflicts of interest.

This article does not contain any studies involving animals performed by any of the authors.

This article does not contain any studies involving human participants performed by any of the authors.

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Translated by I. Gordon

Abbreviations: dNTP—deoxyribonucleoside triphosphate; FBS—fetal bovine serum; LAMP—loop-mediated isothermal amplification; LN—lymph node; OSNA—one-step nucleic acid amplification; PC—prostate cancer.

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Fomicheva, K.A., Makarova, J.A. In Vitro System for the Detection of Prostate Cancer Markers via Loop-Mediated Isothermal Amplification. Appl Biochem Microbiol 56, 881–884 (2020). https://doi.org/10.1134/S0003683820090057

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  • DOI: https://doi.org/10.1134/S0003683820090057

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