(471f) Molecular Combing of Oligonucleotides Induces Secondary Structure Specific Azimuthal Liquid Crystal Orientation

Since its advent in the 1990s, molecular combing of DNA has been exploited for DNA sequencing, gene expression mapping, and pathogen detection. To date, only long (>kbp) DNA strands have been used in these strategies, presumably due to inadequate methods for characterizing the elongation of short oligonucleotides. Previous work in our group using λ-DNA revealed DNA structure specific azimuthal liquid crystal (LC) orientations at near single-molecule sensitivity. In the current work we demonstrate that a similar phenomenon is observed when using short (<20 nucleotides) oligonucleotides. Amine-terminated ssDNA was immobilized onto a (3-glycidoxypropyl) trimethoxysilane modified substrate and stretched by introducing a receding meniscus in a controlled direction. We found that immobilized ssDNA induced azimuthal LC orientation with the LC molecules aligned in the stretching direction.  When we introduced a complementary ssDNA molecule to the aqueous phase prior to stretching, hybridization occurred and the resulting azimuthal orientation was rotated 30° from the stretching direction. In order to demonstrate the potential of this approach for biosensing applications, a microfluidic device was also used to introduce the receding meniscus. Correlation was observed between the shape of the receding air-water interface and the azimuthal LC orientation indicating the expected DNA stretching perpendicular to the interface. This is the first demonstration where molecular combing was used to stretch short oligonucleotides on a solid substrate in a uniform direction, thus illustrating the potential for a highly sensitive DNA hybridization detection strategy that can readily be developed into a multiplexed assay.