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Scanning of guanine–guanine mismatches in DNA by synthetic ligands using surface plasmon resonance

Nature Biotechnology volume 19, pages 51–55 (2001)Cite this article

Abstract

Here we have designed and synthesized ligands that specifically bind with high affinity (Kd = 53 nM) to the guanine (G)–guanine mismatch, one of four types of single-nucleotide polymorphism (SNP). Detection of the G-G mismatch was performed by a surface plasmon resonance (SPR) assay using a sensor chip carrying the G-G specific ligand on its surface. The accuracy of the G-G mismatch detection by the SPR sensor was demonstrated by a marked SPR response obtained only for the DNA containing the G-G mismatch. DNAs containing G-A and G-T mismatches, as well as a fully matched duplex, produced only a weak response. Furthermore, this assay was found applicable for the detection of SNP existing in PCR amplification products of a 652-nucleotide sequence of the HSP70-2 gene.

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Figure 1: Molecular design of a ligand that selectively binds to the G-G mismatch.
Figure 2: DNase I footprinting analysis of the complex of ligand 1 and 5′-32P-end-labeled 64-mer DNA duplex containing G-G, G-A, and G-T mismatched sites.
Figure 3: Illustrations of SPR assay by a sensor chip with covalently immobilized ligand 2 on its surface.
Figure 4: Typical response features (sensorgrams) of the specific binding of DNA containing a G-G mismatch to the 2-immobilized SPR sensor.

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Acknowledgements

The authors thank Prof. Yusuke Nakamura, Dr. Toshihiro Tanaka, Dr. Tatsuhiko Tsunoda, and Dr. Yozo Ohnishi of the Institute of Medical Science, University of Tokyo, for generous gift of PCR products of the HSP70-2 gene. We also thank Dr. Junichi Mineno of Takara Shuzo Co., Ltd. for kind support of our experiments. This work was supported in part by a grant-in-aid for creative basic research (genome science) and for scientific research on priority areas (C) “medical genome science” from the Ministry of Education, Science, Sports and Culture of Japan.

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Authors and Affiliations

  1. Department of Synthetic Chemistry and Biological Chemistry Faculty of Engineering, Kyoto University, CREST, Japan Science and Technology Corporation (JST), Kyoto, 606-8501, Japan

    Kazuhiko Nakatani, Shinsuke Sando & Isao Saito

Authors
  1. Kazuhiko Nakatani
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  2. Shinsuke Sando
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  3. Isao Saito
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Correspondence to Kazuhiko Nakatani.

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Nakatani, K., Sando, S. & Saito, I. Scanning of guanine–guanine mismatches in DNA by synthetic ligands using surface plasmon resonance. Nat Biotechnol 19, 51–55 (2001). https://doi.org/10.1038/83505

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