Chemical amplification for in-gel DNA detection

Abstract

We report the use of reversible addition–fragmentation chain transfer (RAFT) polymerization as a highly efficient chemical amplification means to direct visualization of DNA in porous polyacrylamide gel. It is the first time that a dynamic polymer growth on the surface of soft medium is used in signal amplification for DNA detection. In the proof-of-concept experiment, a thin acrylamide gel on a glass microscope slide formed a thin layer of uniformly crosslinked network with porous structures. Oligonucleotides of different sequences were entrapped within the gel at separate spots. Hybridization of complementary DNA detection probes introduced chain transfer agents (CTAs) into the gel via preconjugation to the probes. Surface-initiated polymer growth was prompted on the gel surface and the growth of polymer brushes at the spot where DNA hybridization occurred was monitored using infrared spectroscopy and atomic force microscopy. Visible change in the texture of the porous gel occurred after polymer growth, which offered an attractive detection alternative for in-gel DNA analysis. Compared to the results from traditional ethidium bromide staining, better detection sensitivity and specificity were achieved.