Visual Platform Based On DNA Nano-hooks For General Detection Of MiRNA In Cancer Cells

MicroRNAs(miRNAs),as cancer markers,were used for cancer diagnosis,treatment,and prognosis monitoring.The occurrence and development of cancer are often associated with the abnormal expression of a variety of miRNAs.A certain RNA can be a biomarker of various inflammation,cancer,or other diseases.Only evaluating the level of a certain miRNA cannot provide an effective reference for clinicians to diagnose a specific disease.The development of a high-throughput and rapid detection platform for multiple miRNAs not only provide multiple indicators and comprehensive information,but also improve the accuracy,which is of great significance for disease diagnosis,treatment,and prognosis monitoring.In this thesis,visual biosensing platforms were developed via the hybridization chain reaction(HCR)to generate DNA nano-hooks,which were then combined with gold nanoparticles(AuNPs)and green synthetic carbon dots(CDs)as follows:1 A colorimetric sensing platform for HCR signal amplification combined with AuNPs was developed for the detection of multi-miRNAs in tumor cells.In the blank experiment,the 3’ and 5’ ends of the free recognition strand were hybridized with the HCR primer at the same time,triggering the HCR reaction to form a DNA macromolecule covered with 15 polymerized thymine strands(poly(A)15)on the surface,which called DNA nanohook.The surface poly(A)15 of the DNA nanohook adsorbed AuNPs,resulting in the aggregation of AuNPs.The solution turned from wine red to colorless.When the target miRNA was introduced,the recognizing strand hybridized with the miRNA to form a rigid dsDNA,preventing the HCR primer from being opened and inhibiting the HCR reaction.The AuNPs were protected as monodisperse,and the solution remained wine red.The detection range of this method is 2.0 × 102-2.0 × 104 HepG2 cells,and the linear equation is:A=0.6338 × IgC-0.9327(C is the number of HepG2 cells),which is expected to become a general detection platform for evaluating the levels of multiple miRNAs.2 A colorimetric/fluorescent dual-output sensing platform via AuNPs-binding CDs was proposed for the detection of miR-222,miR-122,and miR-21 in tumor cells.We obtained CD from food raw materials by green synthesis method.The CD combined with AuNPs can simultaneously output visible light and fluorescence signals.Taking advantage of this capability,we constructed a colorimetric/fluorescence dual-output sensing platform based on DNA nanohooks.The DNA strands self-assembled according to the design in the blank experiment,forming aggregated DNA nanohooks and causing AuNP aggregation.When CD was added,the fluorescence intensity remained high.The introduction of the target miRNA resulted in the homogeneous distribution of AuNPs in the short-chain DNA.The addition of CD resulted in its binding to AuNPs,the red solution turned purple,and the fluorescence of CD was subsequently quenched.Compared with the single-output method,this colorimetric/fluorescent dual-output sensing platform can avoid the influence of background signals and improve the accuracy of measurement,which is expected to be a universal dual output platform for evaluating miR-222,miR-122 and miR-21 in tumor cells.