Detection of fragmented genomic DNA by PCR-free piezoelectric sensing using a denaturation approach

Development of well-automated and miniaturized gene analysis methods is the objective at which research is currently aiming. Recent works report examples of miniaturized PCR devices1 pointing at high-speed PCR. However, a great improvement in DNA sequence analysis would come by direct detection in nonamplified genomic DNA. Biosensors represent an interesting candidate for DNA detection. In particular, several DNA-based sensors have been reported. Most of the work was applied to PCR-amplified samples, and only few works, operating directly with genomic DNA, appeared in the literature with different detection principles. In this paper, a piezoelectric sensor for direct detection of sequences in nonamplified genomic DNA is reported. The system relies on realtime and label-free detection of the hybridization reaction between an immobilized probe (25-mer) and the complementary sequence in solution. The DNA probe is immobilized on the gold electrodes of 10 MHz quartz crystals. Genomic DNA was extracted from the plant, Nicotiana glauca, used as a model system. The target sequence was a portion of the promoter region (35S, present in various genetically modified organisms (GMOs), used as a marker for GMO screening.