Functional Nucleic Acid-based Label-free Fluorescent Biosensors And Logic Gates

Nucleic acid is the carrier of genetic information which widely presents in all animal and plant cells and in microorganisms. The discovery of functional nucleic acids raises our understanding of nucleic acids to a new level and expands the research and use of nucleic acid molecules. Functional DNAs are widely utilized in separation and analysis, clinical diagnosis, targeted drug delivery and the sensor construct due to its unique advantages. In this paper, the detection methods of ions with high sensitivity and selectivity and logic gate systems with the responses to different ions were designed based on label-free functional nucleic acids.Firstly, a signal-on Pb2+ detection method was developed based on the fact that Pb2+ has a significant stabilizing effect for the formation of G-quadruplex structure. The oligonucleotide known as AGRO100 was in random-coil in the absence of Pb2+, while it formed G-quadruplex structure in the presence of Pb2+. G-quadruplex, but not the random-coil structure, can bind to NMM to resulting in the increased fluorescence. Because the formation of the G-quadruplex structure was dependent on the concentration of Pb2+, the fluorescence intensity increased upon increasing the Pb2+ concentration. The fluorescence intensity(excitation/emission maxima, ca. 399/610 nm) of NMM was proportional to the concentration of Pb2+ over the range 0- 1000 n M, with a linear correlation of R2 = 0.9949. The AGRO100-NMM probe provided limit of detection of 3 n M for Pb2+. This approach allows the determination of the concentrations of Pb2+ with simplicity, high selectivity and high sensitivity.Secondly, we have developed a Tb3+-DNA complex-based fluorescence probes for highly selective and sensitive detection of Ag+ and cysteine. It is reported that the fluorescence of Tb3+ can be sensitized in DNA sequence and structure dependent manner. According to circular dichroism and fluorescence spectroscopy data, we identified the sequence of DNA which sensitized the fluorescence of Tb3+. The fluorescence intensity of Tb3+ was proportional to the concentration of Ag+ over the range 0- 2.5 μM(R2 = 0.9984). The Tb3+-DNA probe provided limit of detection of 0.1 μM for Ag+. The sensor has the very high selectivity and can be used for Ag+ detection in complicated samples. The probe can achieve high specific and sensitive detection of cysteine with limit of detection of 50 n M when detecting cysteine based on Ag+ chelating to cysteine.Finally, a variety of logic gates with several ions as inputs and the fluorescence of NMM as output were constructed based on two special DNA structures, DNA triplex and G-quadruplex structures. Construction of an AND logic gate was achieved with H+ and K+ as two inputs. In the K+ containing buffer solution, construction of an INHIBIT logic gate was done with H+ and Ag+ as two inputs. Also, the high sensitive detection of Ag+ could be realized by the INHIBIT logic gates with the limit of detection of 50 n M. In the acidic buffer solution containing K+, NOR logic gate was constructed with Cu2+ and Ag+ as two inputs.