The Effect of Interfacial Design on the Electrochemical Detection of DNA and MicroRNA Using Methylene Blue at Low-Density DNA Films

Herein, we study the effect of interfacial design, specifically the orientation of a DNA probe strand, the length of the diluent and the type of target (DNA vs. microRNA), on the electrochemical response from methylene blue (MB) at low-density DNA films. The probe DNA interfaces are formed from mixed self-assembled monolayers of single-stranded DNA (ss-DNA) and a hydroxyl terminated alkanethiol diluents on gold surfaces. Variation in the current from the DNA-MB complex after hybridization with complementary and single adenine-cytosine mismatch DNA target is shown to depend on whether the 3' or 5' end is tethered to the electrode and the length of the diluent (6-mercapto-1-hexanol vs. 9-mercapto-1-nonanol). The sensors show better mismatch discrimination of single base pairs when both the duplexes are tethered from the 5' end to the electrode and with a longer-chain (9-mercapto-1-nonanol) diluent. Of particular note is our demonstration that the DNA/microRNA duplex exhibits greater single adenine-cytosine mismatch discrimination when compared to the corresponding DNA/DNA duplex. This study contributes to the development of selective DNA and microRNA biosensors.