National Changhua University of Education Institutional Repository : Item 987654321/1551
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题名: Fluorescence Detection of Single-Nucleotide Polymorphisms Using a Universal Molecular Beacon
作者: Lin, Y.-W.;Huang, C.-C.;Chang, H. -T.
贡献者: 化學系
日期: 2008
上传时间: 2010-11-10T06:48:18Z
摘要: We present a simple and novel assay—employing a universal molecular beacon (MB) in the presence of Hg2+—for the detection of single nucleotide polymorphisms (SNPs) based on Hg2+–DNA complexes
inducing a conformational change in the MB. The MB (T7-MB) contains a 19-mer loop and a stem of a pair of seven thymidine (T) bases, a carboxyfluorescein (FAM) unit at the 5’-end, and a 4-([4-(dimethylamino) phenyl]azo)benzoic acid (DABCYL) unit at the
3’-end. Upon formation of Hg2+–T7-MB complexes through T–Hg2+–T bonding, the conformation of T7-MB changes from a random coil to a folded structure, leading to a decreased distance between the FAM and DABCYL units and, hence, increased efficiency of fluorescence resonance energy transfer (FRET) between the FAM and DABCYL units, resulting in decreased fluorescence intensity of the MB. In the presence of complementary DNA, doublestranded DNA complexes form (instead of the Hg2+–T7-MB complexes), with FRET between the FAM and DABCYL units occurring to a lesser extent than in the folded structure. Under the optimal
conditions (20 nM T7-MB, 20mM NaCl, 1.0 kM Hg2+, 5.0mM phosphate buffer solution, pH 7.4), the linear plot of the fluorescence intensity against the concentration of perfectly matched DNA was linear over the range 2–30 nM (R2 = 0.991), with a limit of detection of 0.5 nM at a signal-to-noise ratio of 3. This new probe provides higher selectivity toward DNA than that exhibited by conventional MBs.
關聯: Nucleic Acids Res., 36, e123
显示于类别:[化學系] 期刊論文

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