Dissertation
Dissertation > Biological Sciences > Molecular Biology > Molecular Genetics

The Applications of Gold Nanoparticle in Single-base Mutation Detection

Author GuoKunPeng
Tutor MoZhiHong
School Chongqing University
Course Analytical Chemistry
Keywords Gold nanoparticle double-stranded probes single-base mutation colorimetric detection fluorescence detection magnetic microparticle ligase
CLC Q75
Type Master's thesis
Year 2008
Downloads 126
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Many pathogenic and genetic diseases are associated with changes in the sequence of particular genes. Among these changes, the point mutation, i.e. single nucleotide polymorphisms (SNP), are the most abundant form of genetic variation. Achieving early, accurate, simple and rapid identification to these single-base mutations is of particular importance for the pathogeny and early therapy of corresponding diseases. Several new detection methods for SNP have been developed in the presented paper.A novel colorimetric point mutation detection method was developed by combininng the distance-dependent optical properties of gold nanoparticles with the high specificity of strand displacement. Unmodified gold nanoparticles were used as reported groups. The probes consist of two complementary oligonucleotides of different length labeled with double thiol group. A perfectly matched target and single base mutation target can be completely discrimination taking advantage of their different ability to displace double-stranded probes. This new method is rapid, simple, low cost, and 1 pmol of target produces color changes observable without instrumentation.Single- and double-stranded oligonucleotides have different propensities to adsorb on gold nanoparticles in colloidal solution. Gold nanoparticle is well known for its higher quenching efficiency than any other organic acceptor molecules in fluorescence resonance energy transfer(FRET). In this thesis, we introduce gold nanoparticle as fluorescence quencher to construct a new kind of fluorescence double-stranded probes.The proposed duplex nucleic acid nanoprobe is composed of two complementary oligonucleotides of different lengths. The shorter signal strand is labeled with a fluorophore (FAM) at the 3’-end. A perfectly matched target and single base mutation target can be completely discrimination taking advantage of their different ability to displace fluorescence double-stranded probes.A novel colorimetric point mutation detection method was developed by combininng the efficient separation of magnetic microparticle with the high-fidelity DNA ligase. A ligase reaction that generates the ligation between perfectly matched probes while no ligation occurred between mismatched ones. The interference of the impurities can easily be removed simply by taking away the magnetic field, so the signal can be amplified greatly. This method can be realized the high sensitive detection of DNA target with single base mutation.

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