Study Of Several DNA Sensors Based On Nanoparticle Label

Deoxyribonucleic acid (DNA) is an important hereditary material.The mutation of gene is closely ralated to genetic diseases. Therefore, it isimportant to detect specific DNA sequence in gene expression analysis,gene mutation detection and disease diagnosis. With the development ofbiotechnology, nano-materials have brought new opportunities forbioanalysis technology. For instance, the magnetic separation techniquebased on magnetic nanoparticles has been widely used for separation andpurification of DNA, proteins and cells. Comparing with organicfluorescent dyes, quantum dots have many advantages in optical propertiesthat they provide a new direction to resolve the problems of organicfluorescent probes. Now, gold nanoparticles have attacted more and moreattention in DNA sensors due to their easy preparation, goodbiocompatibility and unique optical properties. This thesis integrated theadvantages of various nano-materials to construct new DNA sensor, andthe main contents include:(1) Amplified fluorescence detection of DNA based on magneticseparation A novel nanoparticles-based fluorescence detection methodtaking advantage of magnetic separation and amplified fluorescencedetection was developed. This DNA sensor relies on a"sandwich"hybridization strategy with two kinds of thiol-modified oligonucleotideprobes immobilized on gold nanoparticles. With the seperation ofmagnetic nanoparticles, the amplified DNA signals raised frombio-bar-code DNA were detected using a chip-based fluorescencedetection method. The result showed that the detection limit of target DNAprobes was 1 pM. Complementary and mismatched sequences wereclearly distinguished, and the ratio of the background-subtractedfluorescence values for complementary and single-base mismatchedoligonucleotide was 2.12:1.(2) A novel DNA detection method based on magnetic separationand fluorescence resonance energy transfer (FRET) of quantum dotsA new type of DNA biosensor was exploited combining magneticseperation of magnetic nanoparticles with FRET of quantum dots. The Fe₃O₄@SiO₂@CdTe complex was prepared by coating green QDs on thesurface of Fe₃O₄@SiO₂ nanoparticles. DNA probes bound toFe₃O₄@SiO₂@CdTe complex hybridized with target DNAs labeled byorange QDs. The results show that the magnetic/fluorescentFe₃O₄@SiO₂@CdTe complex was prepared successfully. The complex issuper-paramagnetic and high fluorescent. Compared with that of thecontrol, the fluorescent signal of target DNA was enhanced clearly due tothe FRET between green and orange QDs. The complementary DNA andmismatched DNA can be distinguished also.(3) DNA detection based on gold nanorods A DNA detectionsystem based on gold nanorod was constructed by taking the advantage ofthe sensitive optical feature of gold nanorods. Firstly, two kinds ofthiol-DNA probes were assembled on the surface of gold nanorods, then asandwich hybridization was carried out by adding target DNA. Afterhybridization, the target DNA can be detected according to the shift ofsurface plasmon resonance peak of gold nanorods. The aspect ratio ofprepared gold nanorods was 2.7. The detection sensitivity tocomplementary DNA reached 1 pM. Complementary and mismatchedsequences can be distinguished effectively. This method is simple andconvenient, which can be used to detect other biomolecules.