Highly sensitive and accurate detection of Vibrio cholera O1 OmpW gene by fluorescence DNA biosensor based on gold and magnetic nanoparticles
2018
Narmani, Asghar | Kamali, Mehdi | Amini, Bahram | Kooshki, Hamid | Amini, Ali | Hasani, Leila
Ultrasensitive DNA based fluorescent assay is used for the determination of Vibrio cholera O1 OmpW gene. OmpW as a surface adherence protein plays a major role in virulence of bacteria with high gene expression. In the present study, two nanoparticles including gold nanoparticles (AuNPs) and magnetic nanoparticles (MNPs) were used. MNPs were functionalized with the first target-specific DNA probe1, which can recognize one end of the target DNA sequence (t DNA) and the AuNPs were functionalized with the second target-specific DNA FAM-probe 2 which can recognize the other end of the target gene. After hybridization of probes with the target DNA, the sandwich structure was formed as MNPs-probe1-target DNA-FAM-probe2-AuNPs by applying the magnetic field to separate the sandwich complex from no hybridized materials. Dithiothreitol (DTT) was used to decompose the sandwich structures and release the probes from target DNA and nanoparticles. Then, fluorescence emission of the released FAM probe 2 on surface AuNPs was detected by fluorescence spectrophotometry. The resulting nanomaterial system demonstrated excellent sensitivity (LOD=2.34ngmL−1) with good linear relationship (R2=0.9989) between target DNA and great selectivity. In conclusion, the present method has a good potential as a rapid detector of bacteria and other infection.
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