Construction And Study Of Novel Fluorescence Sensor For Hepatocellar Carcinoma-related MicroRNAs And DNA

Nucleic acids are a natural choice as recognition elements due to their intrinsic molecular base pairing ability,the detection of nucleic acids is important in biological studies,clinical diagnostics,and biodefense applications.In recent years,the developed fluorescence method has the advantages of easy operation and high sensitivity,thus the methods for detection is one of the most widely used methods.Starting from the point of view of the simple and cost-effective,several novel fluorescent biosensors were constructed based on 2-aminopurine(2-AP)and G-quadruplex.The details are summarized as follows:1.Double-strand displacement biosensor and quencher-free fluorescence strategy for rapid detection of micro RNAThe sensor is designed with an 2-AP probe and a predesigned c DNA.The c DNA partly complement the 2-AP probe.When the target micro RNA(miRNA)is added,the c DNA can be competed off from the c DNA\2-AP probe duplex,thereby forming a c DNA\RNA heteroduplex.The free 2-AP probe induces an increase in the fluorescent signal.A limit of detection of 5 n M and a wide linear range from 5 to 1000 n M(R2 = 0.9971)are achieved by this assay.The rapid detection strategy can be accomplished within 2 h without expensive nanoparticles and complicated instruments for the whole procedure,thus,offering a significant potential for clinical application2.Simply and facilely simultaneous detection of two hepatocellar carcinoma-related micro RNAs using a single-labeled fluorescent probeThe simultaneous detection of two important miRNAs can potentially evaluate pathological states.The probe has a 2-AP as a fluorophore,which is G-quadruplex single-stranded DNA.The probe can partly complement c DNA-1 and c DNA-2.c DNA-1 and c DNA-2 are complementary strands of miRNA-26 a and miRNA-122,respectively.When the target miRNAs are added simultaneously,these c DNA(c DNA-1 and c DNA-2)can be competed off from the c DNA\probe duplex to form a c DNA\RNA heteroduplex.The probe is released and the fluorescent signal is increased.The detection limits of miRNA-26 a and miRNA-122 are both 2.5 n M,and their wide linear which ranges from 2.5 to 500 n M are achieved using the assay.This method has potential practical applications.3.Sensitive label-free fluorescence detection of DNA based on G-quadruplex hairpin DNA.A facile label-free and cost-effective sensing method is developed for the accurate and sensitive fluorescent detection of DNA.The G-quadruplex sequence in H1 is originally locked.Then,when target DNA is present,the hairpin structure of H1 can be unfolded.The H2 hybridizes with the unfolded H1 and displaces the target DNA.Finally,the displaced target DNA again hybridizes with the H1 and initiates cycle amplification.Thus,the numerous G-quadruplexs at the H1 ends are formed,resulting in the fluorescent enhancement after binding with NMM.Lower background signal improves the accuracy of assay.The enzyme-free method can detect down to 40 p M and a linear range of 3 orders of magnitude.Moreover,this strategy has an ability to discriminate the target DNA from even single-base mismatched or other sequence.The proposed method has potential applications in DNA-based molecular diagnostics.