Analytical Sciences

Synthesis of Circular Double-Stranded DNA Having Single-Stranded Recognition Sequence as Molecular-Physical Probe for Nucleic Acid Hybridization Detection Based on Atomic Force Microscopy Imaging

Koji NAKANO,* Hideshi MATSUNAGA,* Masaharu MURATA,** Nobuaki SOH,* and Toshihiko Imato*

*Department of Applied Chemistry, Faculty of Engineering, Kyushu University, 744 Motooka, Nishi, Fukuoka 819-0395, Japan
**Department of Advanced Medical Initiatives, Faculty of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi, Fukuoka 812-8582, Japan

A new class of DNA probes having a mechanically detectable tag is reported. The DNA probe, which consists of a single-stranded recognition sequence and a double-stranded circular DNA entity, was prepared by polymerase reaction. M13mp18 single strand and a 32mer oligodeoxynucleotide whose 5′-end is decorated with the recognition sequence were used in combination as template and primer, respectively. We have successfully demonstrated that the DNA probe is useful for bioanalytical purposes: by deliberately attaching target DNA molecules onto Au(111) substrates and by mechanically reading out the tag-entity using a high-resolution microscopy including atomic force microscopy, visualization/detection of the individual target/probe DNA conjugate was possible simply yet straightforwardly. The present DNA probe can be characterized as a 100%-nucleic acid product material. It is simply available by one-pod synthesis. A surface topology parameter, image roughness, has witnessed its importance as a quantitative analysis index with particular usability in the present visualization/detection method.

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