Conformational change detection of DNA with the fluorogenic reagent of o-phthalaldehyde-beta-mercaptoethanol.
o-Phthalaldehyde-beta-mercaptoethanol (OPAME) as a fluorogenic reagent has been found wide applications in the detection of amino acids based on its reaction with primary amino groups. In this contribution, we report our new findings concerning the reactions of OPAME with single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA), respectively. It has been found that ssDNA can react with OPAME easily as a result of giving rise to strong fluorescence emissions, while dsDNA, prepared by hybridizing ssDNA with its complementary target prior to the reaction, displays inert chemical activity and gives out weak fluorescence emission. Mechanism investigations have shown that the reaction activity between OPAME and DNA depends on the amino groups that are related to the conformation of uncoiled and exposed extent of DNA structure, and thus the inert chemical activity of dsDNA results from screening of the dsDNA bases in the interior of the double strands. Therefore, we could design a way to detect conformation change of DNA with OPAME and further develop a novel, simple label-free sequence detection method for complementary and single-base mismatched ssDNA in the hybridization of DNA.
Long
Yun Fei
YF
College of Chemistry and Chemical Engineering, Southwest University, CQKL-LRTA, Chongqing, China.
Liao
Qie Gen
QG
Huang
Cheng Zhi
CZ
Ling
Jian
J
Li
Yuan Fang
YF
D000588:Amines / Q000737:chemistry; D004247:DNA / Q000737:chemistry*; D004277:DNA, Single-Stranded / Q000737:chemistry; D005456:Fluorescent Dyes; D013058:Mass Spectrometry; D008623:Mercaptoethanol / Q000737:chemistry*; D009690:Nucleic Acid Conformation; D009693:Nucleic Acid Hybridization; D013050:Spectrometry, Fluorescence; D013055:Spectrophotometry, Infrared; D013056:Spectrophotometry, Ultraviolet; D009764:o-Phthalaldehyde / Q000737:chemistry*
D000588:Amines; D004277:DNA, Single-Stranded; D005456:Fluorescent Dyes; D008623:Mercaptoethanol; D009764:o-Phthalaldehyde; D004247:DNA