AFM-imaging diagnosis method for single nucleotide polymorphism using molecular beacon DNA as an intramolecular ligation template of target DNA and a viewable indicator

Hisao Yoshinaga, Koji Nakano, Nobuaki Soh, Toshihiko Imato

    Research output: Contribution to journalArticlepeer-review

    4 Citations (Scopus)

    Abstract

    An AFM-imaging-based method for single nucleotide polymorphism (SNP) analysis is described. A stem-loop-forming 34-mer oligonucleotide (p34s) was designed. P34s contains the complementary sequence for K-ras (5'-GGT GGC-3', t6G), one of the human oncogenes, at the 5'-end for target-recognition and five successive phosphorothioate linkages in the loop. The functional probe, either alone or hybridized with target DNA (p34s/t6G), relaxed upon treatment with "opener" DNA. The template/target DNA interstrand hybridization product is covalently connected by ligase if the correct target is used, but not hybridized species including mismatches. With these results, developed was a solid-phase SNP assay by transferring an aliquot of the product onto an Au(111) substrate for self-assembly, followed by AFM imaging. Clear contrasts that allow the detection of SNPs, were observed for the ligated and non-ligated species representing the loop-to-linear conformational change. Simple statistical surface-roughness analysis determined the lowest concentration of the sample to be 5 × 10 -10 M, whose necessary sample quantity was 5 fmol. 2012

    Original languageEnglish
    Pages (from-to)939-945
    Number of pages7
    Journalanalytical sciences
    Volume28
    Issue number10
    DOIs
    Publication statusPublished - 2012

    All Science Journal Classification (ASJC) codes

    • Analytical Chemistry

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