| Summary: | 碩士 === 中原大學 === 生物科技研究所 === 102 === An improvement of the detection of environmental pathogens in water is very important because it requires a rapid detection of contamination sources and inhibition of contaminant expansion. Traditional methods usually use selective medium to culture and measure microorganisms. However, it has some disadvantage in its difficulty of quantification and lengthy incubate time (always 2 to 3 days). Also, there are over a thousand species of micro-organisms present in water and not all species can be detected using the traditional method. The quantum dot platform is consisted of a microparticle binding oligonucleotide and designed probes (internal/reporter probe) coated with quantum dot. In our study, this system could be stable and demonstrate highly specificity and sensitivity. As followed specificity and sensitivity assay, the standard curve could reveal that R2 ratio was 0.9. In addition, the limitation assay could detect Pseudomonas spp. at least 10 bacteria and 0.001 ng/μL Escherichia coli target DNA in water sample. Therefore, an application of a novel detecting technology is needed to resolve these problems. In order to increase specificity and sensitivity efficiency, a quantum-dot detection approach developed a high throughput methodology based on previous study in 2012, called as a DNA fingerprinting system. The Fingerprinting system was established different length of fragments by restriction enzyme cutting site, called as Terminal Restriction Fragment Length Polymorphism (T-RFLP), to diversify sizes for evaluating specificity. Interestingly, our research had a time-efficient and an easier detection system that avoid the use of the ineffective traditional-chip methods and PCR amplification. Our technique obtained (1) highly specific, (2) highly sensitive, (3) accurate quantification, and (4) a DNA fingerprinting analysis of environmental pathogens in water samples.
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