Photochemical control of the polymerase chain reaction has been achieved through the incorporation of light-triggered nucleotides into DNA.Photochemical activation enables the precise spatial and temporal regulation of chemical and biological function. This is typically achieved through the installation of photochemically removable protecting groups (caging groups) on the molecule of interest, often a biological macromolecule. These caging groups are then removed in a spatially and temporally restricted fashion through irradiation with UV light (decaging), leading to activation of the molecule under study. 1 Caging has been employed in the photochemical regulation of several processes, e.g. enzymatic activity, 2 gene expression, 3 as well as DNA and RNA function. 4,5 Here, we report on the photochemical regulation of the polymerase chain reaction (PCR). PCR was developed in 1983, and is employed in the in vitro isolation and exponential amplification of specific DNA sequences. 6 By utilizing thermophilic DNA polymerases with specifically designed DNA primers, extremely small amounts of DNA can be rapidly enriched to substantial quantities. In the few years since its discovery, PCR has revolutionized the field of molecular biology, facilitating genome sequencing, genetic disease diagnosis, and genetic fingerprinting. 7 We expect that the photoregulation of PCR will afford an additional level of control over this important technique.Recently, we developed a novel caging group for N-heterocyclic molecules 8 and applied it to the specific caging of a thymidine nucleotide on its heterocyclic base. 4 The corresponding caged phosphoramidite was incorporated into DNA oligomers using standard DNA synthesis equipment and protocols. 4 Installation of the sterically demanding caging group in conjunction with the disruption of an N-H bond critical for Watson-Crick base pairing allowed for the attenuation of catalytic activity of a DNAzyme. Brief irradiation with UV light of 365 nm (25 W, handheld UV lamp) removes the caging group and generates the regular DNA oligomer (Scheme 1). In order to apply this approach to the photochemical regulation of PCR, we first investigated the effect of one or multiple caging groups on the hybridization to a complementary DNA strand. The DNA oligomers P1-P7, consisting of 19 nucleotides, a typical length for PCR primers, and containing 0-4 caged thymidines have been synthesized (Table 1). These primers were then analyzed for their annealing and melting properties in the presence of a complementary oligonucleotide.Melting curves were measured on a BioRad MyiQ RT-PCR thermocycler by conducting a sequence of 3 heating and cooling Cycles (1 μM of both primer and complementary DNA

Cre recombinase catalyzes DNA exchange between two conserved lox recognition sites. The enzyme has extensive biological application, from basic cloning to engineering knock-out and knock-in organisms. Widespread use of Cre is due to its simplicity and effectiveness, but the enzyme and the recombination event remain difficult to control with high precision. To obtain such control we report the installation of a light-responsive o-nitrobenzyl caging group directly in the catalytic site of Cre, inhibiting its activity. Prior to irradiation, caged Cre is completely inactive, as demonstrated both in vitro and in mammalian cell culture. Exposure to non-damaging UVA light removes the caging group and restores recombinase activity. Tight spatio-temporal control over DNA recombination is thereby achieved.

Four-arm, star-shaped poly(D,L-lactide) (PDLLA) oligomers of controlled molar mass and narrow molar mass distribution were successfully synthesized by use of an ethoxylated pentaerythritol initiator. Derivatization of the terminal hydroxyl groups with either methacrylic anhydride (MAAH) or 2-isocyanatoethyl methacrylate (IEM) to yield PDLLA-M (M = methacrylate end group) and PDLLA-UM (UM = urethane methacrylate end group), respectively, was monitored by in situ Fourier transform infrared (FTIR) spectroscopy. Photo-cross-linking of the functional oligomers yielded networks with high gel contents (>95%). The glass transition temperature (T(g)) of these networks was strongly dependent on prepolymer molar mass, and networks based on low molar mass precursors were more rigid than the networks obtained from higher molar mass oligomers. The tensile strength (TS) and Young's modulus of the PDLLA-M samples, approximately 7 and 17 MPa, respectively, were significantly lower than the values of 19 MPa (TS) and 113-354 MPa (Young's modulus) for the PDLLA-UM samples. The introduction of terminal hydrogen-bonding sites that were adjacent to the photo-cross-linking site resulted in higher performance poly(lactide)-based bioadhesives.

The effect of microwave irradiation on DNA/DNA hybridization has been studied under controlled power and temperature conditions. It was discovered that microwave irradiation led to the melting of double-stranded deoxyoligonucleotides well below their thermal melting temperature and independent of the length of the deoxyoligonucleotides. These observations indicate a specific interaction of microwaves with DNA, and have important implications in the chemical or enzymatic processing of DNA under microwave heating.