title: DNA-binding small-ligand-immobilized surface plasmon resonance biosensor for detecting thymine-related single-nucleotide polymorphisms.
abstract: A surface plasmon resonance (SPR) biosensor that carries DNA-binding small ligands has been developed for the detection of single-nucleotide polymorphisms (SNPs). 3,5-Diaminopyrazine derivatives, with a hydrogen-bonding profile fully complementary to the thymine base, were utilized as recognition elements on the sensor surface, and a target single-stranded DNA sequence was hybridized with a DNA probe containing an abasic site to place this site opposite a nucleobase to be detected. In a continuous flow of sample solutions buffered to pH 6.4 (0.25 M NaCl), the 3,5-diaminopyrazine-based SPR sensor can detect an orphan nucleobase in the duplex with a clear selectivity for thymine over cytosine, guanine, and adenine (5'-GTT GGA GCT GXG GGC GTA GGC-3'/3'-CAA CCT CGA CNC CCG CAT CCG-5'; X=abasic site, N=target nucleobase G, C, A, or T). The SPR response was linear in the concentration range 10-100 nM. Allele discrimination is possible based on the combination of different binding surfaces in a flow cell of the SPR system, which is demonstrated for the analysis of the thymine/cytosine mutation present in 63-meric polymerase chain reaction (PCR) amplification products (Ha-ras gene, codon 12, antisense strand). Comparison with a bulk assay based on 3,5-diaminopyrazine/DNA binding shows that the immobilization of 3,5-diaminopyrazine derivatives on the SPR sensor allows more sensitive detection of the target DNA sequence, and binding selectivity can be tuned by controlling the salt concentration of sample solutions. These features of the DNA-binding small-molecule-immobilized SPR sensor are discussed as a basis for the design of SPR biosensors for SNP genotyping.
authors.affiliation: Department of Chemistry, Graduate School of Science, Tohoku University, Aoba-ku, Sendai 980-8578, Japan.
authors.forename: Sara
authors.initials: S
authors.lastname: Miura
authors.forename: Seiichi
authors.initials: S
authors.lastname: Nishizawa
authors.forename: Akinori
authors.initials: A
authors.lastname: Suzuki
authors.forename: Yukiko
authors.initials: Y
authors.lastname: Fujimoto
authors.forename: Katsuya
authors.initials: K
authors.lastname: Ono
authors.forename: Qiang
authors.initials: Q
authors.lastname: Gao
authors.forename: Norio
authors.initials: N
authors.lastname: Teramae
mesh_terms: D001483:Base Sequence; D015374:Biosensing Techniques / Q000379:methods; D004247:DNA / Q000737:chemistry*; D015342:DNA Probes / Q000737:chemistry*; D008024:Ligands; D015394:Molecular Structure; D020641:Polymorphism, Single Nucleotide; D020349:Surface Plasmon Resonance / Q000379:methods; D013941:Thymine / Q000737:chemistry*
chemical_list: D015342:DNA Probes; D008024:Ligands; D004247:DNA; D013941:Thymine