Development Of Electrochemical DNA Sensor And Application In The Detection Of Staphylococcus Aureus And Enterotoxin

Staphylococcus aureus(S. aureus) is an very common pathogens cause of food poisoning, so the research on rapid detecting technology of S. aureus is particularly important. Nowadays, the detection methed of S. aureus is mainly depend on traditional detection, but it have many shortcomings such as time-consuming, difficult to quantitative detection and so on. In recent years, the electrochemical DNA sensors as a new DNA testing technology because of its rapid, sensitive and convenient features, developed rapidly in the field of food safety and environmental testing. This paper is aiming to build a electrochemical DNA sensor with high sensitivity and low limit of detection by virtue of excellent sensor materials, supply a more rapidly, convenient and efficient detection method for S. aureus.The study firstly builds a detection electrode on the basis of multi-walled carbon nanotubes and thionine joint modified glassy carbon electrode. The DNA sensor selects the specific nucleotide sequence of nuc gene as a DNA probe by the hybridization indicator of methylene blue, and makes the phosphate group modified on the5’end of DNA sequence, in order to combined with the amino of the thionine through a covalent bond. In this way the probe sequence could be connected to the electrode to detect the target sequence of the S. aureus. And this experiment is also makes a optimization of the proportioning between multi-walled carbon nanotubes and thionine, the enrichment time of MB, the temperature and time of target sequence hybridization. Finally, The detection limit of the specific fragment in the nuc gene of the S. aureus could reach2×10-11mol/L.Then, A new electrode electrochemical DNA sensor which based on AuNPs/p-ABA/MWNTs-COOH modified glassy carbon is builded, the specific DNA probe is incorporated into the electrode fabricated with gold nanoparticles in the self-assembly way, and the signal of doxorubicin which acted as electrical activity hybridization indicator was detected by the method of DPV. The detection limit of the specific fragment in the nuc gene of the S. aureus could reach1×10-12mol/L. This research lay a foundation for the application of the sensing technology in high efficient and real-time detection of S. aureus.Finally, we adopt the AuNPs/p-ABA/MWNTs-COOH modified electrode as the detection electrode with low limit of detection and high sensitivity, which combined separately with the specific sequence of the nuc-gene and the specific sequence which control the production of SEA to detect the S. aureus. Studies have shown that the electrochemical DNA sensor we have built could realize the detection for the S. aureus in the samples and was able to identify the Enterotoxigenic Staphylococcus aureus A.