The Research On Spectrum Methods For Detecting Sequence-specific Dna Of Genetically Modified Cattle

With the development process of genetically modified organisms promoting to the commercialization, the issues of safety assessments and safety managements on GMO were also attracted much attention. In the safety assessments, the most important task is to test the transgenic samples. Numerous analytical methods have been developed for reliable determination of the presence or absence of genetically modified organisms in various products. Most of the methods rely on the two strategies either based on detecting the inserted foreign DNA or the novel protein that is specifically expressed in transgenic organisms. DNA based testing methods with rapidity and high accuracy can be widely applied. However, it is still a challenge to detect long oligonucleotides specifically. The fluorescent probe has triggered great interests for its high specificity based on fluorescence resonance energy transfer. Recently, gold nanoparticles possess distinct physical and chemical attributes that make them excellent scaffolds for the fabrication of novel chemical and biological sensors. To solve the problems on the detection of GMO,we developed three new methods for detecting long sequence-specific oligonucletiodes of genetically modified cattle based on fluorescent probe and gold nanoparticles optical molecular probes. Those spectrometric methods were based on probe hybridization and combined with fluorophore-based techniques and nanoanalysis techniques. The main contents and conclusions of this paper are as follows:1. Two fluorescent probes(FITC molecular beacon and Texas Red molecular beacon)were fabricated by long oligonucleotides. They hybridized to adjacent regions on the same target resulting in dual fluorescence resonance energy transfer. This method had many desirable features incluing rapid detection, high specificity and accuracy, which were suitable for detecting long sequence-specific nucleic acid fragments of transgenic cattle.2. Gold nanoparticle probes in response to targets specifically were synthesized by derivatizing gold nanoparticle solution with hairpin structure long alkanethiol-modified oligonucleotides. Capture probes and gold nanoparticle probes co-hybridized with targets,which was the basis of the colorimetric sensors. This nanoparticle-based three-component sandwich hybridization assay realized it that the colorimetric detection of PCR products of transgenic cattle could be visualized with the naked eye.3. Gold nanoparticle probes in response to targets specifically were synthesized by derivatizing gold nanoparticle solution with hairpin structure long alkanethiol-modified oligonucleotides. The assay relied on the observation of greatly enhanced resonance light scattering that originated from the aggregation of oligonucleotide-functionalized gold nanoparticles directed by the target DNA. This was a simple, cost-effective, highly sensitive detection of long sequence-specific nucleic acid fragments of transgenic cattle.The above methods made it possible for specifically detecting long sequence-specific DNA, which met the needs of different detection environment, and provided a new idea for GMO detection.