TTP plc (TTP), a leading independent technology and product development company, has successfully demonstrated a reverse transcriptase-free RNA detection method. The approach can be used with a standard qPCR master mix and a universal hydrolysis probe to produce concentration-dependent qPCR data without the need for a reverse transcriptase.
Instead of being used as a template to generate cDNA, the target RNA is bound by oligonucleotides with two distinct sections. The first of these sections binds to the target while the second, in the presence of the specific RNA target, forms part of a complex that also includes a universal primer and a synthetic target sequence. When the complex is stable, which is only when the RNA target is present, the synthetic target sequence can be amplified to produce a PCR product. From this point onwards, the reaction becomes a simple qPCR reaction with hydrolysis probe detection.
The two significant costs in standard RNA analysis systems - the reverse transcriptase enzyme and the hydrolysis probe – are either eliminated or made universal. While this method does require the production of target-specific oligonucleotides, these are generally very low cost, particularly when used for large numbers of reactions. This combination could see RNA detection become more universally available and accessible to even the most cost-sensitive markets.
While there have been attempts to drive down costs of RNA analysis to some degree, the need for a reverse transcriptase has remained a barrier to truly low-cost tests. Reverse transcription remains essential for some applications, e.g. for monitoring the changes to a viral sequence over time, but in a lot of cases the only questions being asked are whether a virus or RNA sequence is present and at approximately what concentration. In such a scenario this simpler, cheaper system devised by TTP has the potential to remove cost barriers and lead to more universal implementation of RNA detection and quantification.
TTP will be exhibiting at the 71st AACC Annual Scientific Meeting & Clinical Lab Expo, at booth #3383 in California, 4–8 August 2019.