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Electrochemical Analysis of Sybr Green I and its Interaction with DNA
Dudová, Zdenka ; Pivoňková, Hana ; Havran, Luděk ; Fojta, Miroslav
There are used many methods such real-time PCR and electrophoresis in molecular biology and biochemistry which are based on utilization of a fluorescent dye Sybr Green I (SG) for selective detection of double stranded (ds) DNA and its quantification. SG is a planar molecule (see Fig. 1) that intercalates into the DNA double helix and simultaneously can bind into minor groove of dsDNA. In our work we focused on electrochemical behavior of SG on a pyrolytic graphite electrode (PGE) using square-wave voltammetry (SWV). The voltammetric method was used in measurements of SG interactions with DNA at the PGE.
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Electrodeposited Coatings at ITO/FTO Enhancing their Applicability in Electroanalysis
Daňhel, Aleš ; Fojta, Miroslav
Recent possibilities of modification of transparent electrodes based on indium-tin oxide and/or fluorine doped tin oxide by electrodeposited coatings enhancing their electrochemical applicability in electroanalysis are discussed in this work. Ordered mesoporous silica and/or chemically functionalized mesoporous silica thin films may serve, among the common utilization in solid phase extraction, as protective layer of the electrode surface against its fouling and/or as molecular sieve permeable for small molecules and blocking large ones (DNA, proteins, etc.). Furthermore an electrodeposition of organic modifiers and/or metal (nano) particles inside of the ordered silica nanochanels together with direct electrodeposition of selected metal (nano) particles onto selected conductive transparent supports are presented here as well.
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Electrochemical Analysis of Chemically Modified DNA at Gold Electrodes
Vidláková, Pavlína ; Havran, Luděk ; Fojta, Miroslav
DNA labeling is used to increase sensitivity of its electrochemical detection. Covalently bound markers based on a complex of osmium tetroxide with 22-bipyridine(Os,bipy) is one of those frequently applied. Os bipy reacts with pyrimidines (mainly thymines) in single stranded regions of DNA, therefore it represents a useful marker for the analysis of DNA structure or labeling of hybridization probes . Another possibility of DNA labeling is the incorporation of chemically modified dNTPs into DNA molecules with using primer extension. DNA labeling in connection with voltammetry at gold electrodes has application potential in development of sensors for studying of DNA structure and interaction.
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Synthetic Biology Introduces New Base Pairs to Analyze
Špaček, Jan
An unnatural base pair is composed of two nucleobase analogues which specifically Pair only with each other similarly as natural DNA pairs do. Goal of the search for new base pair is expansion of genetic alphabet with new codons containing unnatural bases. Expanded genetic alphabet am, other things allows simultaneous site specific incorporation of non-standard amino acids into proteins Recently unnatural bases forming pairs vie hydrophobic interactions, which can be replicated by prokaryotic cells with efficiency and mutation rate similar to natural base pairs, have been shown. Synthetic biology provides new opportunities for application of electrochemical analysis.
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Electrochemical Analysis of Carrageenans. The Effect of Acetate Concentration
Kos, S.S. ; Paleček, Emil
kappa-, iota-, and lambda-carrageenans are polysaccharides (PSs) which produce electrocatalytic chronopotentiometric peak H-ps in acetate buffer (pH 5.2) on the hanging mercury drop electrode (HMDE). Here we show that this peak (E-p1.8V) increases with the acetate concentration Using a. c. voltammetry we found that the appearance and the observed increase of peak H-PS is accompanied by a decrease of capacitive current below the background elecnolyte around1.8 V, suggesting strong adsorption of the negatively charged PS at the negatively charged HMDE. Compared to carrageenany dextran did not produce peak H-PS and did not strongly adsorb at negatively charged electrode.
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Constant Current Chronopotentiometry as a Method for Detection of Singlet Oxygen Protein Damage
Ostatná, Veronika ; Vargová, Veronika ; Černocká, Hana ; Paleček, Emil
Proteins are one of the major targets for oxidative damage in the cell.Indirect non-radical oxidation of the protein via formation and subsequent reaction with single oxygen (O-1(2))is one of the major processes.In this work we studied the single oxygen(O-1(2))-mediated oxidation of bovine serum albumin (BSA) by constant current chronopotentiometry in combination with mercury electrode.Our chronopotentiometric data show that photo-oxidized BSA was more susceptible to the electric field-induced denaturation than non-oxidized native BSA. Our method utilizing intrinsic electrochemical signal of proteins provides a sensitive detection methods for minor damages in various proteins.
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New Ligands for Modification of DNA by Osmium Tetroxide Reagents
Havran, Luděk ; Zenka, Martin ; Fojta, Miroslav
Besides DNA intrinsic electroactivity, modification of DNA by redox active tags find wide use in electrochemical analysis of DNA interactions and damage. Application of these tags can improve specificity of the analysis. One from utilized labels in this field are complexes of osmium tetroxide with nitrogen ligands, which produce with DNA electroactive covalent adducts. These complexes preferentially react with thymine in single strand DNA. Type of used ligand influence electrochemical behavior forming adducts. In this contribution will be presented results which are acquired by testing of set a new ligands based of quinone and substituted 1, 10-phenanthroline.
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