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Electroanalytical study of proflavine intercalation in 5-methyl or inosine-containing amplicons

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Abstract

Amplicons corresponding to the GC-rich p53 exon 5 and its analogues, synthesized by substituting 60% of cytosine by 5-methyl-cytosine, or 60% of guanosine by inosine and GC-poor p53 exon 6 were synthesized and investigated electrochemically, in the presence and absence of proflavine, by differential pulse voltammetry (DPV). Incorporation of base analogues and the thermal stability of the resulting amplicons were tested in the presence of a fluorescent probe (Sybr–Green). Peak current at 1.0 V was lower for methylated than for unmethylated PCR amplicons and was similarly affected by proflavine intercalation. In contrast, considerable peak current differences were observed in the presence of proflavine for unmodified exon 5 v.s. exon 6 or inosine-containing amplicons. Thermal analysis verified the expected shifts in melting temperature (T m) due to the base analogue incorporation and GC-content variations. In conclusion, methylated and unmethylated PCR amplicons could be distinguished in model DNA systems using differential pulse voltammetry (DPV) and use of proflavine could serve as an electrochemical probe for identifying different DNA conformations.

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Acknowledgments

This paper is part of the 03ED835 research project, implemented within the framework of the “Reinforcement Programme of Human Research Manpower” (PENED) and co-financed by National and Community Funds (25% from the Greek Ministry of Development-General Secretariat of Research and Technology and 75% from E.U.-European Social Fund).

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Authors and Affiliations

  1. Laboratory of Analytical Chemistry, Department of Chemistry, Aristotle University of Thessaloniki, 541 24, Thessaloniki, Greece

    Despina K. Alexiadou, Andrea K. Ioannou, Anastasios N. Voulgaropoulos & Stella Th. Girousi

  2. Laboratory of Biochemistry, Department of Medicine, Aristotle University of Thessaloniki, 541 24, Thessaloniki, Greece

    Sofia A. Kouidou-Andreou

Authors
  1. Despina K. Alexiadou
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  2. Andrea K. Ioannou
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  3. Sofia A. Kouidou-Andreou
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  4. Anastasios N. Voulgaropoulos
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  5. Stella Th. Girousi
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Correspondence to Stella Th. Girousi.

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Alexiadou, D.K., Ioannou, A.K., Kouidou-Andreou, S.A. et al. Electroanalytical study of proflavine intercalation in 5-methyl or inosine-containing amplicons. Anal Bioanal Chem 392, 533–539 (2008). https://doi.org/10.1007/s00216-008-2285-4

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  • DOI: https://doi.org/10.1007/s00216-008-2285-4

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