Development of an Interaction Assay between Single-Stranded Nucleic Acids Trapped with Silica Particles and Fluorescent Compounds
Biopolymers are easily denatured by heating, a change in pH or chemical substances when they are immobilized on a substrate. To prevent denaturation of biopolymers, we developed a method to trap a polynucleotide on a substrate by hydrogen bonding using silica particles with surfaces modified by amin...
| Main Authors: | , |
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| Format: | Article |
| Language: | English |
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MDPI AG
2012-09-01
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| Series: | Journal of Functional Biomaterials |
| Subjects: | |
| Online Access: | http://www.mdpi.com/2079-4983/3/3/601 |
| Summary: | Biopolymers are easily denatured by heating, a change in pH or chemical substances when they are immobilized on a substrate. To prevent denaturation of biopolymers, we developed a method to trap a polynucleotide on a substrate by hydrogen bonding using silica particles with surfaces modified by aminoalkyl chains ([A-AM silane]/SiO<sub>2</sub>). [A-AM silane]/SiO<sub>2</sub> was synthesized by silane coupling reaction of N-2-(aminoethyl)-3-aminopropyltrimethoxysilane (A-AM silane) with SiO<sub>2</sub> particles with a diameter of 5 μm at 100 °C for 20 min. The surface chemical structure of [A-AM silane]/SiO<sub>2</sub> was characterized by Fourier transform infrared spectroscopy and molecular orbital calculations. The surface of the silica particles was modified with A-AM silane and primary amine groups were formed. [A-AM silane]/SiO<sub>2</sub> was trapped with single-stranded nucleic acids [(Poly-X; X = A (adenine), G (guanine) and C (cytosine)] in PBS solution at 37 °C for 1 h. The single-stranded nucleic acids were trapped on the surface of the [A-AM silane]/SiO<sub>2</sub> by hydrogen bonding to form conjugated materials. The resulting complexes were further conjugated by derivatives of acridine orange (AO) as fluorescent labels under the same conditions to form [AO:Poly-X:A-AM silane]/SiO<sub>2</sub> complexes. Changes in the fluorescence intensity of these complexes originating from interactions between the single-stranded nucleic acid and aromatic compounds were also evaluated. The change in intensity displayed the order [AO: Poly-G: A-AM silane]/SiO<sub>2</sub> > [AO:Poly-A:A-AM silane]/SiO<sub>2</sub> >> [AO:Poly-C:A-AM silane]/SiO<sub>2</sub>. This suggests that the single-stranded nucleic acids conjugated with aminoalkyl chains on the surfaces of SiO<sub>2</sub> particles and the change in fluorescence intensity reflected the molecular interaction between AO and the nucleic-acid base in a polynucleotide. |
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| ISSN: | 2079-4983 |