(113c) Morpholino Microarrays

A principal challenge in microarray-based analysis of nucleic acids is to detect hybridization between probe molecules on the array with complementary target strands from solution against a background of competing base-pairing interactions (e.g. probe-probe associations, double-stranded targets). Synthetic DNA analogs, whose hybridization with nucleic acid targets can exhibit qualitatively different dependence on experimental conditions than that between two nucleic acid strands, provide a route for improving selectivity of on-array hybridization relative to other possible base-pairing associations. In this work, uncharged DNA analogues called morpholinos (MOs) are investigated as microarray probes instead of DNA. Morpholino microarrays are fabricated by contact printing on aldehyde slides. On these substrates, in addition to covalent immobilization MOs were found to efficiently immobilize through physical adsorption. Physically adsorbed probes could be removed by washes with surfactant solutions. Comparison of solution and surface hybridization thermodynamics reveals how salt concentration, temperature, and selection of probe type (DNA or MO) impact selectivity for the desired reaction on the microarray support relative to other possible base-driven associations. For example, interrogation of double-stranded DNA targets with morpholino microarrays revealed a hybridization maximum at intermediate salt concentrations, and comparison of surface and solution hybridization free energies indicates influence of probe/surface interactions. These observations are discussed in the context of the molecular organization of MO and DNA probe films.