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NBD‐Based Green Fluorescent Ligands for Typing of Thymine‐Related SNPs by Using an Abasic Site‐Containing Probe DNA
Author(s) -
Thiagarajan Viruthachalam,
Rajendran Arivazhagan,
Satake Hiroyuki,
Nishizawa Seiichi,
Teramae Norio
Publication year - 2010
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.200900530
Subject(s) - ap site , thymine , dna , fluorescence , chemistry , microbiology and biotechnology , computational biology , genetics , biology , combinatorial chemistry , dna repair , physics , quantum mechanics
The binding behavior of green fluorescent ligands, derivatives of 7‐nitrobenzo‐2‐oxa‐1,3‐diazole (NBD), with DNA duplexes containing an abasic (AP) site is studied by thermal denaturation and fluorescence experiments. Among NBD derivatives, N 1 ‐(7‐nitrobenzo[ c ][1,2,5]oxadiazol‐4‐yl)propane‐1,3‐diamine (NBD‐NH 2 ) is found to bind selectively to the thymine base opposite an AP site in a DNA duplex with a binding affinity of 1.52×10 6 M −1 . From molecular modeling studies, it is suggested that the NBD moiety binds to thymine at the AP site and a protonated amino group tethered to the NBD moiety interacts with the guanine base flanking the AP site. Green fluorescent NBD‐NH 2 is successfully applied for simultaneous G>T genotyping of PCR amplification products in a single cuvette in combination with a blue fluorescent ligand, 2‐amino‐6,7‐dimethyl‐4‐hydroxypteridine (diMe‐pteridine).