Bulletin of the Korean Chemical Society

Korean Chemical Society (대한화학회)
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A Linear Beacon System Featuring an Internal Deoxyguanine Quencher Allows Highly Selective Detection of Single Base Mismatches

  • Lee, Young-Ae (Department of Chemistry, Kyungpook National University) ;
  • Hwang, Gil-Tae (Department of Chemistry, Kyungpook National University)
  • Received : 2010.05.12
  • Accepted : 2010.05.27
  • Published : 2010.07.20
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Abstract

The fluorescence intensity of a single-stranded oligonucleotide containing a fluorene-labeled deoxyuridine $(U^{Fl})$ unit increases by only 1.5-fold upon formation of its perfectly matched duplex. To increase the fluorescence signal during hybridization, we positioned a quencher strand containing a deoxyguanine (dG) nucleobase, functioning as an internal quencher, opposite to the $U^{Fl}$ unit to reduce the intrinsic fluorescence upon hybridization with a probe. From an investigation of the optimal length of the quencher strand and the effect of the neighboring base sequence, we found that a short strand (five-nucleotide) containing all natural nucleotides and dG as an internal quencher was effective at reducing the intrinsic fluorescence of a linear beacon; it also exhibited high total discrimination factors for the formation of perfectly matched and single base-mismatched duplexes. Such assays that function based on clear changes in fluorescence in response to single-base nucleotide mutations would be useful tools for accelerating diagnoses related to various diseases.

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