RNA Fluorescent Probes: Design,Synthesis And Their Application In Cells And Living Systems

Ribonucleic acid?RNA?is a biological macromolecule composed of ribonucleotides,which mainly affects the genetic coding,regulation and expression in living cells.RNA,as an important signal molecule,transmits genetic information from DNA to proteins and also participates in gene expression and signal transduction processes of living systems.RNA also controls gene expression by altering its structure.Therefore,achieving visualization of RNA is very important to study the role of RNA in regulating cell metabolism.Small-molecule fluorescent probes are indispensable tool for biological imaging.At present,a variety of small-molecule fluorescent probes have been used for targeted detection of biological molecules in living organisms.At the same time,there is a certain relationship between nucleic acids and other cellular components?such as certain biological species and organelles?.Small-molecule fluorescent probe can be used as a new approach to understand their positions,functions,and relationships in physiology and pathology.RNA fluorescent probes have become powerful and versatile tools for studying the function of biological RNA.Compared with RNA fluorescent probes designed based on the structure of proteins and oligonucleotides,small-molecule RNA fluorescent probes are distinguished on account of their unique advantages?such as high spatial resolution,good membrane permeability,and excellent photophysical properties?.However,there is only one commercial fluorescent probe?SYTO RNA Select?for RNA analysis at present,so it is of great significance to design and synthesize small molecular fluorescent probes that can specifically detect RNA.Based on the current research situation,the paper designs and synthesizes three small-molecule fluorescent probes that can recognize RNA in biological systems,and further explains their applications through experiments.In chapter two,a novel fluorescent probe TP-MIVC based on carbazole group was designed.The probe can simultaneously detect RNA and H₂S in various biological systems with different fluorescent signals.In the optical experiment,TP-MIVC showed weak fluorescence in the aqueous solution.With the increase of RNA,the fluorescence intensity increased by 12 times at 625 nm,and in the presence of HS^-,the fluorescence intensity increased significantly at 550 nm.TP-MIVC can detect H₂S in the blue channel and recognize RNA in the red channel of cell imaging.TP-MIVC can simultaneously detect RNA and H₂S in zebrafish with two sets of different fluorescent signals.In addition,the probe TP-MIVC was able to distinguish normal mice and tumor mice by imaging RNA for the first time.In chapter three,a small molecule probe EPI-RS was constructed that based on indole groups for imaging RNA and SO₂ with two sets of different fluorescence signals in living systems.EPI-RS used"turn on"and"turn off"fluorescent signals to image RNA and SO₂ in biological imaging,respectively.Due to the unique V-shaped structure of the probe,it can be seen that the fluorescence increases with the addition of RNA during the spectral test.RNase digestion experiments confirmed that the probe can recognize intracellular RNA,and the probe mainly concentrated in the nucleoli.In addition,EPI-RS visualized RNA in mouse liver tissue and applied to zebrafish and mouse.In chapter four,a new conjugate system was constructed and the pyrrole single ion salt MR-IDE was designed and synthesized.Using this ionic salt as a new type of fluorescent platform,it recognized RNA based on a special structure and introduced mitochondrial site to identify mtRNA.Colocalization experiments revealed that MR-IDE was distributed in the cytoplasmic mitochondria,and the fluorescence intensity in the nucleoli was significantly higher than that in the nucleus.RNase digestion experiments had further demonstrated that the probe could recognize the RNA of cancer cells and macrophages.MR-IDE has the advantages of high selectivity,light stability,low cytotoxicity,and good cell membrane permeability.