The principle behind using diphenylamine reagent to identify DNA
Release date:
2025-04-03
The principle behind using diphenylamine reagent to identify DNA is that, when heated under acidic conditions, the deoxyribose in DNA is converted into ω-hydroxy-γ-ketopentanal, which then reacts with diphenylamine to produce a blue-colored compound.
Specific reaction mechanism
Acidic conditions and glycosidic bond cleavage
In DNA's purine nucleotides, deoxyribose is linked to the purine base via a glycosidic bond. When heated in an acidic (H⁺) environment, the glycosidic bond breaks, releasing free deoxyribose.
Deoxyribose conversion
Under acidic conditions and upon heating, free deoxyribose undergoes dehydration to form the intermediate product ω-hydroxy-γ-ketopentanal.
Color development reaction
ω-Hydroxy-γ-ketopentanal combines with the diphenylamine reagent, producing a blue compound via a redox reaction. This color change serves as a distinctive marker for the presence of DNA.
Experimental condition requirements
Boiling water bath heating: Accelerates the cleavage of glycosidic bonds and the conversion of deoxyribose, enhancing reaction sensitivity. Typically, heating is required for more than 5 minutes.
Cooling
Important Notes
This reaction specifically targets DNA, but its sensitivity is relatively low, requiring a higher concentration of DNA to produce a visible color change.
Reagent preparation requires proportionally mixing Liquid A (acidic environment) and Liquid B (containing diphenylamine) to ensure the accuracy of the reaction system.
This method is widely used in biological experiments, such as the identification steps in DNA extraction protocols.
Jiangsu Feiya Chemical Industry Group Co., Ltd.
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