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A facile plasmonic nanoplatform was developed for rapid and sensitive determination of nucleic acid. Hg 2+ -regulated molecular beacon (MB, hairpin) containing rich thymine (T) bases at both ends is used as the probe. A hairpin structure can be formed in the MB probe due to the strong binding of Hg 2+ to T. However, in the presence of target DNA, the hairpin structure is opened owing to target DNA-specific hybridization with the aptamer. Simultaneously, the opened MB interacts with poly(diallyldimethylammonium chloride) (PDDA) and hinders PDDA-induced aggregation of AuNPs, accompanied by a color change from blue to red and a decrease in absorption ratio (A 620 /A 520 ). Hence, a good linear relationship was observed between the decreased absorption ratio (A 620 /A 520 ) and DNA concentration ranging from 0.02 to 2 nmol/L with a low detection limit of 4.42 pmol/L. Moreover, this nanoplatform has been successfully utilized to discriminate between perfect target and mismatch sequences. More importantly, the bioassay is simple, versatile, rapid, and cost-effective compared with other common methods, which holds great promise for clinical diagnosis and biomedical application. Graphical abstract.

Background: Recently, some research results showed that the circulating DNA in serum or plasma had potential for the molecular diagnosis and prognosis of certain cancers. Several methods have been employed for the quantification of circulating DNA. However, the circulating DNA levels obtained by various methods exhibited considerable differences. Additionally, these methods were labor-extensive and time-consuming, and not suitable for the quantification of circulating DNA in numerous samples due to the use of commercial DNA extraction kits for the purification of circulating DNA. We presented a new method for the quantification of circulating DNA in sera by capillary zone electrophoresis (CZE) with laser-induced fluorescence detection (LIF)., Methods: In the present work, we want to make comparison between CZE-LIF assay and real time PCR for the quantification of circulating DNA levels. Linearity, intra and inter variability of two methods were evaluated., Results: The intra and inter variability of circulating DNA quantification by real-time PCR were 7.3% and 14.92%, respectively. In CZE assay the intra and inter variability were 4.19% and 6.91%, respectively. The R.S.D. values of the same coated capillary and different coated capillaries were 5.14% and 9.02%, respectively. Our data showed that the circulating DNA levels obtained by two methods had a good correlation. Moreover, we further confirmed that blood samples collection, serum preparation and other treatment procedures had a significant impact on the DNA levels in sera., Conclusion: Our data further illustrated that CZE-LIF is a simple, rapid and sensitive method for the quantification of circulating DNA in human sera, and well suitable for the analysis of a large number of samples in clinical diagnosis.

EvaGreen is a new DNA intercalating dye successfully used in quantitative real-time PCR. In the present work, we firstly apply EvaGreen to the analysis of dsDNA by CE with LIF detection. Comparisons of EvaGreen dye with the commonly used dyes SYBR Green I and SYBR Gold were preformed in dsDNA analysis by CE. The linear range of dsDNA using EvaGreen was slightly wider than that using SYBR Gold and SYBR Green I, and the detection limits of dsDNA were not significantly different for the three dyes. Good separations of dsDNA fragments were obtained using the three dyes. Reproducibility of migration time and the peak area of dsDNA fragments with EvaGreen were better than those for SYBR Green I and SYBR Gold. The RSD values were 0.24-0.27% for migration time and 3.45-7.59% for peak area within the same day, 1.35-1.63% for migration time and 6.72-12.05% for peak area for three days. Our data demonstrated that EvaGreen is well suited for the dsDNA analysis by CE with LIF detection.

Hot-start (HS) effects were investigated in pfu-based polymerase chain reaction (PCR), when water-soluble CdTe quantum dots (QDs) were introduced in the PCR system. The HS effects were demonstrated by the higher amplicon yields and excellent suppression of non-specific amplification after pre-incubation of PCR mix with QDs between 35°C and 56°C. DNA targets were well amplified even after PCR mixture was pre-incubated 1 h at 50°C. Importantly, the effects of QDs nanoparticles could be reversed by increasing the pfu polymerase concentration, suggesting that there was an interaction between QDs and pfu DNA polymerase. Moreover, control experiment indicated that HS effect is not primarily due to the reduced pfu polymerase concentration resulted from the above interaction. Fluorescence correlation spectroscopy (FCS), a single molecule detection method, was used to investigate the possible mechanism of HS PCR with QDs. Preliminary FCS results suggested that CdTe QDs may directly interact with pfu DNA polymerase, rather than other components in the PCR system. Furthermore, results demonstrated that the interaction between QDs and pfu resulted in a reduction in pfu polymerase concentration. This study provided a good start to investigate potential implications of QDs in other key molecular biology techniques. [ABSTRACT FROM PUBLISHER]