New methods for simultaneous multiplex RNA and DNA amplification and detection

Researchers from the Department of Biotechnology and Systems Biology have published new methods for one- or multiplex amplification and detection of RNA or DNA targets.

The detection of pathogenic microorganisms present in food, feed, plant, and other samples is important for providing safe food as well as for preventing the spread of microbes.

With a goal of simultaneous detection of pathogens with genomes in RNA or DNA form, researchers from the Department of Biotechnology and Systems Biology, have developed a new NAIMA procedure, suitable for simultaneous multiplex amplification of RNA and DNA targets, coupled with the detection on ArrayTubes. The method was demonstrated to be very sensitive and specific for the detection of two economically important quarantine plant pathogens of potato, the potato spindle tuber viroid (RNA target) and Ralstonia solanacearum (DNA target). Because of its isothermal amplification and simple detection equipment, the method is also applicable for on-site analyses. NAIMA can be used in any domain where there is the need to detect RNA and DNA targets simultaneously.

Additionally, for the detection of abovementioned pathogens, our researchers have developed new loop-mediated isothermal amplification (LAMP) methods, which shorten the time needed for analyses to minimum, because the isolation of nucleic acids is not needed and the amplification takes only about 20 minutes.

All methods were published in scientific journals:

• D. Dobnik, D. Morisset, R. Lenarčič, M. Ravnikar. Simultaneous Detection of RNA and DNA Targets Based on Multiplex Isothermal Amplification. Journal of Agricultural and Food Chemistry, 2014, 62 (13), 2989-2996
• R. Lenarčič, D. Morisset, N. Mehle, R. Ravnikar. Fast real-time detection of Potato spindle tuber viroid by RT-LAMP. Plant Pathology, 2012, 62 (5), 1365-3059
• R. Lenarčič, D. Morisset, M Pirc, P. Llop, M. Ravnikar, T. Dreo. Loop-mediated isothermal amplification of specific endoglucanase gene sequence for detection of the bacterial wilt pathogen Ralstonia solanacearum. PLoS ONE 9(4): e96027