Detection of Specific RNA Targets by Multimerization

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.

Automated summary

In this study, we propose an RNA target detection method based on nucleic acid multimerization. This technique only requires a single DNA polymerase with reverse transcriptase, DNA-dependent DNA polymerase, and strand-displacement activities. Efficient detection of target RNAs through multimerization mechanism was achieved, and the approach was validated using genetic material of the SARS-CoV-2 coronavirus. The multimerization reaction allowed reliable differentiation between SARS-CoV-2 RNA-positive and SARS-CoV-2 negative samples, even in samples subjected to multiple freezing-thawing cycles.