Studies And Applications Of Multifunctional Oriented Nanoprobes For Nucleic Acid Analysis

With optical signal intensity, good stability and excellent optical properties, noble metal nanoparticles have become a new kind of optical probe and have been widely applied in the field of biological imaging, chemical detection, nano optics. Therefore, constructing multifunctional noble nanoprobes with excellent performance based on the integration of unique nanomaterials with nanoassembly and modification, is significant for the development of chemical detection, biological imaging, and intracellular physiological processes studying. By the combination of unique nanomaterials with asymmetric modification technique, we have constructed a series of multifunctional oriented monomer and dimer nanoprobes with good stability and excellent optical properties, which having great potential for multiplexed detection of biomolecules in complex biochemical environment. The details are summarized as follows:1. Construction and study of oriented assembly nanoprobesBased on the combination of unique nanomaterials and the asymmetric modification technique, we construct a series of multifunctional monomer and dimer nano probes. Firstly, we made silane modified glass by ourselves, which was positively charged. Monolayer adsorption of nano particles was prepared on glass surface by electrostatic adsorption. Then we made nano materials with single layer adsorption to prepare the asymmetrically modified nano probe. We also studied the optical properties and stability of the nano probe. Due to the advantage of the asymmetric modification technique, we can not only directionally assemble functional molecules and target molecules on the nanoparticles, but also effectively control the amount of assembled molecules. Compared with the conventional nano probes, such nanoprobes have excellent stability and oriented assembly ability in a complex biological detection system, which enables this nano probe for multiplexed biomolecules detection with high selectivity and sensitivity specificity.Then, we present a methodology for highly sensitive imaging of multiplexed mRNA at molecular level, by using novel oriented probes to construct a turn-on signal generation mechanism based on the plasmon resonance scattering of noble metal nanoparticles. Compared to the conventional optical biosensors, this new strategy possesses higher potential in detecting and imaging multi-mRNA. Firstly, Y-shaped DNA duplex actuation of nanoparticle dimers, which can extremely minimize the interparticle distance while maximizing the plasmon coupling effect, are strong enough for sensitive detection of nucleic acid. Secondly, based on the different scattering characteristics of gold and silver nanoparticles, it can achieve simultaneous detection of multi-biomolecule. With proper linker functionalization, such platform could be extended and served as a general basis to pave a way for imaging various biomolecules in complex biochemical environment,.2. Construction and purification of dimer probesWe constructed a variety of dimers nanoprobes based on DNA hybridization by employing the asymmetrically assembled monomer probes. The centrifugal separation was carried out by using the density gradient centrifugation method. In order to purify the prepared dimer, we take different concentration gradient of iodixanol solution as the concentration gradient solution to make centrifugal separation. By using the density gradient centrifugation method, dimer nanoprobes have been successfully separated and purified from the mixed samples. The purification of dimer provides significant promising in the condition without the ultracentrifuge. This provides significant basis for application of dimmers in biomolecule detection, imaging analysis, etc.