Towards autonomous DNA-based nanodevices

Molecular recognition, programmability, self-assembling capabilites and biocompatibility are unique features of DNA. The basic approach of DNA nanotechnology is to exploit these properties in order to fabricate novel materials and structures on the nanometer scale. This cumulative dissertation deals with three aspects of this young research area: fast analysis, autonomous control of functional structures, and biocompatible autonomous delivery systems for nanoscale objects. 1. At low temperatures and under favorable buffer conditions, two complementary DNA strands will form a double-helical structure in which the bases of the two strands are paired according to the Watson-Crick rules: adenine bases bind with thymine bases, guanine bases with cytosine bases. The melting temperature TM of a DNA duplex is defined as the temperature at which half of the double strands are separated into single strands. The melting temperature can be calculated for DNA strands of known sequences under standard conditions. However, it has to be determined experimentally for strands of unknown sequences and for applications under extreme buffer conditions. A method for fast and reliable determination of DNA melting temperatures has been developed. Stable gradients of the denaturing agent formamide were generated by means of diffusion in a microfluidic setup. Formamide lowers the melting temperature of DNA and a given formamide concentration can be mapped to a corresponding virtual temperature along the formamide gradient. Differences in the length of complementary sequences of only one nucleotide as well as a single nucleotide mismatch can be detected with this method, which is of great interest for the detection of sequence mutations or variations such as single nucleotide polymorphisms (SNPs). 2. Knowledge of the stability of DNA duplexes is also of great importance for the construction of DNA-based nanostructures and devices. Conformational changes occuring in artificially generated DNA structures can be used to produce motion on the nanometer scale. Usually, DNA devices are driven by the manual addition of fuel molecules or by the periodic variation of buffer conditions. One prominent example of such a conformational change is the formation of the so-called i-motif, which is a folded four-stranded DNA structure characterized by noncanonical hemiprotonated cytosine-cytosine base-pairs. In order to achieve controlled autonomous motion, the oscillating pH-value of a chemical oscillator has been employed to drive the i-motif periodically through its conformational states. The experiments were conducted with the DNA switch in solution and attached to a solid substrate and constitute the first example of DNA-based devices driven autonomously by a chemical non-equilibrium reaction. 3. Finally, a DNA-crosslinked and switchable polyacrylamide hydrogel is introduced, which is used to trap and release fluorescent colloidal quantum dots in response to externally applied programmable DNA signal strands. Trapping and release of the nanoparticles is demonstrated by studying their diffusion properties using single molecule fluorescence microscopy, single particle tracking and fluorescence correlation spectroscopy. Due to the biocompatibility of the polymerized acrylamide and the crosslinking DNA strands, such