Highly Sensitive and Robust Linear Probe for Detection of mRNA in Cells

Abstract

As temless linear probe was designed that robustly detects mRNAi nc ells with high sensitivity.T he probe is modified at some positions with base surrogates prepared from d-threoninol, with anthraquinone moieties near the 5'-and 3'termini, and with perylene moieties.E ven in cell lysate that involves various proteins and enzymes,b ackground emission was very low.W hen the probe was hybridized with RNA, chromophores are intercalated between the base pairs,resulting in ar emarkable light-up signal. The signal-to-background ratio was as high as 1600 under our standardbuffer conditions. In the HeLa cell lysate,t he linear probe had sufficient signalto-background ratio (S/B = 40) for reliable mRNAd etection. No degradation was observed after a24hincubation in HeLa cell lysate.Incells,aprobe designed to target DsRed resulted in distinct blue fluorescence only in cells transfected with plasmid encoding DsRed;n of luorescence was observed in control cells.