title: Recognition of a C-C mismatch in a DNA duplex using a fluorescent small molecule with application for "off-on" discrimination of C/G mutation.
abstract: The fluorescent small molecule 2-amino-7-methyl-1,8-naphthyridine (AMND) can selectively bind to a cytosine (C) at a C-C mismatch in double-stranded DNA (dsDNA). The interactions between AMND and C-C mismatch-containing dsDNA were investigated by measuring ultraviolet (UV) absorption as a function of temperature to obtain melting curves as well as circular dichroism and fluorescence spectra. Results show that AMND strongly stabilizes C-C mismatch-containing dsDNA, whereas fully matched duplexes are not stabilized under the same conditions. The fluorescence of AMND was efficiently quenched when it was bound to a C-C mismatch in dsDNA. Binding constants (K(11)), obtained by fluorescence titration, were 1.2 × 10(5) M(-1). Although sensing functions depend on the sequences flanking the mismatch site, the change in AMND fluorescence intensity can be utilized to detect the C-C mismatch-containing dsDNA. Accordingly, discrimination of the C/G mutation in the model sequence (PGR gene rs1255998) was achieved by visualizing fluorescence of AMND. A probe DNA molecule was designed to contain a C opposite the C/G base in the target DNA, and this probe was used to hybridize the target DNA. The fluorescence of AMND was "on" for a C-G match, while the fluorescence was "off" for a C-C mismatch. This assay is simple and does not require DNA labeling.
authors.affiliation: Key Laboratory of Applied Surface and Colloid Chemistry, Ministry of Education, School of Chemistry and Materials Science, Shaanxi Normal University, Xi'an 710062, China.
authors.forename: Liangliang
authors.initials: L
authors.lastname: Hu
authors.forename: Yunxiang
authors.initials: Y
authors.lastname: Wang
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.forename: Qiang
authors.initials: Q
authors.lastname: Gao
authors.forename: Honglan
authors.initials: H
authors.lastname: Qi
authors.forename: Chengxiao
authors.initials: C
authors.lastname: Zhang
mesh_terms: D020137:Base Pair Mismatch; D002942:Circular Dichroism / Q000379:methods; D003596:Cytosine / Q000737:chemistry*; D004247:DNA / Q000032:analysis / Q000737:chemistry*; D004252:DNA Mutational Analysis / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry*; D006147:Guanine / Q000737:chemistry*; D009287:Naphthyridines / Q000737:chemistry; D009691:Nucleic Acid Denaturation; D020641:Polymorphism, Single Nucleotide; D013056:Spectrophotometry, Ultraviolet
chemical_list: C520210:2-amino-7-methyl-1,8-naphthyridine; D005456:Fluorescent Dyes; D009287:Naphthyridines; D006147:Guanine; D003596:Cytosine; D004247:DNA