A colorimetric confirmation method for DNA amplification in PCR and its application to the detection of Giardia lamblia cysts.
A simple and quick colorimetric method for confirming DNA amplification in polymerase chain reactions (PCR) is described and has been applied to the amplification of Giardia lamblia DNA. This method detects the release of pyrophosphate based on the competition between 1, 10-phenanthroline and pyrophosphate complexing with ferrous ion. When 1, 10-phenanthroline complexed with Fe2+ is added to the finished PCR solution, depending on whether or not the DNA was amplified, the mixture is, respectively, either bleached or red. The color changed optimally for 20-30 min at 60-80 degrees C, and the result could be determined by detecting an absorbancy change at 510 nm or a color change discernible to the naked eye. The extent of change in absorbance was proportional to the amount of pyrophosphate produced.
Lee
Kisay
K
Department of Environmental Engineering and Biotechnology, Myongji University, Yongin, Kyongki, 449-728, Korea. kisay@mju.ac.kr
Kim
Kyungju
K
D000818:Animals; D003124:Colorimetry / Q000379:methods*; D004247:DNA / Q000032:analysis* / Q000096:biosynthesis*; D016829:Giardia lamblia / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013696:Temperature
D004247:DNA