May 31, 2014 - bacterial targets based on a simultaneously amplified DNAzyme. Youngung .... of PCR amplified products of pathogenic bacteria that cause.
Microchim Acta (2014) 181:1965–1971 DOI 10.1007/s00604-014-1297-3
ORIGINAL PAPER
Colorimetric detection of PCR products of DNA from pathogenic bacterial targets based on a simultaneously amplified DNAzyme Youngung Seok & Ju-Young Byun & Hyoyoung Mun & Min-Gon Kim
Received: 22 January 2014 / Accepted: 17 May 2014 / Published online: 31 May 2014 # Springer-Verlag Wien 2014
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.
