Preparation Of Quantum Dots Conjugated Molecular Beacon Probe And Application For DNA Detection

A kind of novel hairpin molecular beacons probe (MBP) was presented in this paper. Inorganic fluorescent CdTe quantum dots (QDs) and quenchers (for example DABCYL) were connected by single-stranded DNA (ssDNA) to get hairpin structure of MBP. It was confirmed that there was a part of overlap between emission spectrum of CdTe QDs and absorption spectrum of quenchers. Therefore, the fluorescence resonance energy transfer (FRET) could occur between CdTe QDs donors and DABCYL quencher acceptors when their distance was close enough (about 1-10 nm). Moreover, the distance between donors and acceptors were controlled by the hybridization of DNA, and the change of distance of donors and acceptors would lead to the change of fluorescence intensity of MBP. Accordingly, a complementary DNA with ssDNA could be detected by this fluorescent probe system.The CdTe QDs with 2.5 nm in diameter were synthesized with mercaptopropyl acetic acid (HS-CH2COOH) as stabilize agent in water, which exhibited a strong, stable and narrow band (full width at half maximum, FWHM was 24 nm) in emission spectrum, and an optimization reaction condition was explored by changing the pH of the reaction system, refluxing time, and so on.According to the principle of FRET, the emission spectrum of CdTe QDs and the absorption spectrum of quenchers were measured respectively to get suitable CdTe QDs as energy donor, and the CdTe QDs were modified by the 5' end of DNA-Quencher in phosphate-buffered solution (PBS). When the 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) as dehydration agent was existed in the system, the carboxyl on the CdTe surfaces could react with amino of DNA. It was indicated that the FRET system was set up successfully when the fluorescence intensity of MBP was decreased extremely after the modification of CdTe QDs.Moreover, the minimum limit of detection and specificity of MBP was studied according to the changes of the fluorescence intensity of the MBP system afterhybridization between MBP and different target DNA. The fluorescence intensity of the MBP system recovered to the maximum after hybridization between the MBP and a complete complementary target DNA, which demonstrated that the FRET structure of MBP was unfolded and the distance between CdTe QDs donors and the quencher acceptors was increased, therefore, the fluorescence intensity of MBP recovered. The minimum limit of detection of MBP system with complementary DNA in this paper was 5.039×10^-9mol/L according to the measures of fluorescence intensity changes after hybridization. When a mismatching base-pair DNA was as target DNA, the fluorescence intensity of MBP system was increased but which was much less than the increase when the complementary DNA was as target. These results indicated that this MBP system not only had an excellent sensitivity and specificity with complementary DNA, but also could detect the base of target DNA was mutant or not.