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Colorimetric detection of PCR products of DNA from pathogenic bacterial targets based on a simultaneously amplified DNAzyme

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Abstract

A novel strategy was devised for colorimetric analysis of the products of the polymerase chain reaction (PCR). The method takes advantage of simultaneous amplification of a horseradish peroxidase-mimicking DNAzyme (HRPzyme) during the PCR process. It is performed using a DNA specific forward primer and a universal reverse primer containing a complementary HRPzyme sequence. The double-strand PCR products, which include the HRPzyme sequence, are treated with a mixture of hemin and TMB (3,3′,5,5′–tetramethylbenzidine) in the presence of hydrogen peroxide. The resulting HRPzyme/hemin complex then promotes a peroxidase mimicking reaction, which produces the blue colored oxidized TMB. This colorimetric method can be more easily performed than previously developed gel based detection procedures and, as a result, can be conveniently applied to the specific and sensitive colorimetric analysis of DNA sequences arising from pathogenic bacteria. The potentially broad applicability of the new method has been demonstrated by its use in the identification of the 16s rDNA of Salmonella Typhimurium.

A novel strategy was devised for simple colorimetric analysis of PCR products with amplification of a horseradish peroxidase-mimicking DNAzyme(HRPzyme). This colorimetric method can be much more easily performed than previously developed gel based detection procedures and potentially broad applicability for other DNA analysis.

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Acknowledgments

This work was financially supported by the R&D Joint Venture Program, NLRL Program (2011-0028915) and New Industry Creation Project through the National Research Foundation of Korea (NRF) funded by the Korean government (Ministry of Science, ICT and Future Planning).

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Authors and Affiliations

  1. Department of Chemistry, School of Physics and Chemistry, Gwangju Institute of Science and Technology (GIST), 261 Cheomdan-gwagiro, Gwangju, 500-712, Republic of Korea

    Youngung Seok, Ju-Young Byun, Hyoyoung Mun & Min-Gon Kim

  2. Advanced Photonics Research Institute, Gwangju Institute of Science and Technology (GIST), 261 Cheomdan-gwagiro, Gwangju, 500-712, Republic of Korea

    Min-Gon Kim

Authors
  1. Youngung Seok
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  2. Ju-Young Byun
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  3. Hyoyoung Mun
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  4. Min-Gon Kim
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Correspondence to Min-Gon Kim.

Additional information

Youngung Seok and Ju-Young Byun contributed equally to this work.

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Seok, Y., Byun, JY., Mun, H. et al. Colorimetric detection of PCR products of DNA from pathogenic bacterial targets based on a simultaneously amplified DNAzyme. Microchim Acta 181, 1965–1971 (2014). https://doi.org/10.1007/s00604-014-1297-3

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  • DOI: https://doi.org/10.1007/s00604-014-1297-3

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