Electrochemically active DNA probes: detection of target DNA sequences at femtomole level by high-performance liquid chromatography with electrochemical detection

Electrochemically active DNA probes were prepared by linking a ferrocene unit with 5'-aminohexyl-terminated oligonucleotides. The DNA sequences of probes 5a, 5b, and 5c were 5'-T12-3', 5'-T20-3', and 5'-TGCAG TTCCG GTGGC TGATC-3', respectively. Probe 5a could form a complex selectively with a single-strand poly(A) and a double-strand DNA fragment containing an A13 sequence and these complexes could be detected at femtomole levels by an electrochemical detector (ECD) on HPLC. The observed ECD response was proportional to the amount of the complex over the range 20-100 fmol. Probe 5c was capable of detecting femtomole levels of a restriction DNA fragment having oncogene v-myc. Moreover, probe 5b was able to detect picogram levels of mRNA taken from rat brain or yeast total cellular RNA. This proves that the electrochemically active DNA probes are useful in analyzing traces of DNA and RNA carrying the complementary sequence.