Preparation And Study Of Nucleic Acid Probe Based On Polystyrene-Gold Nanoparticles Composite Microspheres

Polystyrene(PS)microspheres are often used as the carrier materials in liquid biochips for high-throughput detection of biomolecules because of the micrometer scale size and easy to functional modification.They are.Molecular beacon is the nucleic acid probe with stem-loop structure,which can detect single nucleotide polymorphism because of its better specificity than linear nucleic acid probe.In this paper,polystyrene microspheres were combined with molecular beacon and Au NPs were used to quench the fluorescent substances,replacing the traditional quenching groups of molecular beacon.As a result,a new nucleic acid probe based on polystyrene – gold nanoparticles composite microspheres was developed.The specific research contents are as follows:(1)Monodisperse polystyrene microspheres were prepared by dispersion polymerization.The results showed that the prepared monodisperse microspheres had good monodispersity and a narrow particle size distribution.The surface was smooth,and the diameter was about 2μm.(2)The surface of polystyrene microspheres was coated with a layer of poly dopamine by self-polymerization of dopamine in alkaline solution.The gold nanoparticles were deposited on the surface of the microspheres by the reduction of chloroauric acid in situ by the dopamine.The particle size was about 20 nm.Then,the particle was taken as the nucleus,the reducing agent hydrochloric acid hydroxylamine was used to reduce the chloroauric acid,and the gold nanoparticles were grown.Finally,the composite microspheres with gold nanoparticles were obtained.(3)A molecular beacon with one end thiolated was connected to the surface of polystyrene nanogold by Au-S bond to prepare a nucleic acid probe.The effect of NaCl concentration on the molecular beacon loading on the surface of the microspheres was investigated.The loading of molecular beacon on the surface of the microspheres was up to 40 nmol / g when the NaCl concentration was 0.6 M.(4)The performance of this probe was systematically investigated.The probe had a good specificity,can effectively detect the single nucleotide polymorphism,and detection time was short,about 90% of the fluorescent signal was generated within 10 min.And the target nucleic acid miRNA-21 and TK1 DNA were simultaneously detected by using multifunctional molecular imager and flow cytometer.This study will provide important experimental data for the application of nucleic acid probes based on polystyrene – gold nanoparticles composite microspheres for disease diagnosis.