Detection of Specific RNA Targets by Multimerization

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Assol R. Sakhabutdinova¹, Alexey V. Chemeris¹, and Ravil R. Garafutdinov^1,a*

¹Institute of Biochemistry and Genetics, Ufa Federal Research Center, Russian Academy of Sciences, 450054 Ufa, Bashkortostan, Russia

^* To whom correspondence should be addressed.

Received November 22, 2022; Revised February 26, 2023; Accepted March 9, 2023

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Detection of specific RNA targets via amplification-mediated techniques is widely used in fundamental studies and medicine due to essential role of RNA in transfer of genetic information and development of diseases. Here, we report on an approach for detection of RNA targets based on the particular type of isothermal amplification, namely, reaction of nucleic acid multimerization. The proposed technique requires only a single DNA polymerase possessing reverse transcriptase, DNA-dependent DNA polymerase, and strand-displacement activities. Reaction conditions that lead to efficient detection of the target RNAs through multimerization mechanism were determined. The approach was verified by using genetic material of the SARS-CoV-2 coronavirus as a model viral RNA. Reaction of multimerization allowed to differentiate the SARS-CoV-2 RNA-positive samples from the SARS-CoV-2 negative samples with high reliability. The proposed technique allows detection of RNA even in the samples, which were subjected to multiple freezing-thawing cycles.

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KEY WORDS: ribonucleic acids (RNA), coronavirus SARS-CoV-2, molecular diagnostics, isothermal amplification, multimerization, nearby primers

DOI: 10.1134/S0006297923050103

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