Characterizing the interaction between DNA and GelRed fluorescent stain
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We have performed single-molecule stretching and dynamic light-scattering (DLS) experiments to characterize the interaction between the DNA molecule and the fluorescent stain GelRed. The results from single-molecule stretching show that the persistence length of DNA–GelRed complexes increases as the ligand concentration increases up to a critical concentration, then decreases for higher concentrations. The contour length of the complexes, on the other hand, increases monotonically as a function of GelRed concentration, suggesting that intercalation is the main binding mechanism. To characterize the physical chemistry of the interaction, we used the McGhee–von Hippel binding isotherm to extract physicochemical data for the interaction from the contour length data. Such analysis enabled us to conclude that the GelRed stain is, in fact, a bis-intercalator. In addition, DLS experiments were performed to study the changes of the effective size of the DNA–GelRed complexes, measured as the hydrodynamic radius, as a function of ligand concentration. We observed qualitative agreement between the results obtained from the two techniques by comparing the behavior of the hydrodynamics radius and the radius of gyration, because the latter quantity can be expressed as a function of mechanical properties determined from the stretching experiments.
KeywordsIntercalation Single-molecule stretching Dynamic light scattering Binding isotherm
This work was supported by the Brazilian agencies: Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG) and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq). The DLS experiments were performed in the “Laboratório de Microfluídica e Fluidos Complexos (LMFFC)” of Universidade Federal de Viçosa.
- Biotium (2013) Safety report of gelred and gelgreen. Nucleic acid detection technologies. http://www.biotium.com. Accessed Oct 2014
- Cesconetto EC, Junior FSA, Crisafuli FAP, Mesquita ON, Ramos EB, Rocha MS (2013) DNA interaction with actinomycin d: mechanical measurements reveal the details of the binding data. Phys Chem Chem Phys 15(26):11070–11077Google Scholar
- Daune M (1999) Molecular biophysics, 1st edn. Oxford University Press, OxfordGoogle Scholar
- Fritzsche H, Triebel H, Chaires JB, Dattagupta N, Crothers DM (1982) Interaction of anthracycline antibiotics with bio-polymers. 6. studies on interaction of anthracycline antibiotics and deoxyribonucleic-acid—geometry of intercalation of iremycin and daunomycin. Biochemistry 21(17):3940–3946PubMedCrossRefGoogle Scholar
- Maaloum M, Mullera P, Harlepp S (2013) DNA–intercalator interactions: structural and physical analysis using atomic force microscopy in solution. Soft Matter 9:11233Google Scholar
- Reis LA, Ramos EB, Rocha MS (2013) DNA interaction with diaminobenzidine studied with optical tweezers and dynamic light scattering. J Phys Chem B 117(46):14345–14350Google Scholar
- Rocha MS (2009) Modeling the entropic structural transition of dna complexes formed with intercalating drugs. Phys Biol 6:036013Google Scholar
- Rocha MS, Ferreira MC, Mesquita ON (2007) Transition on the entropic elasticity of DNA induced by intercalating molecules. J Chem Phys 127(10), Art. 105108 Google Scholar
- Rocha MS, Lúcio AD, Alexandre SS, Nunes RW, Mesquita ON (2009) DNA-psoralen: single-molecule experiments and first principles calculations. Appl Phys Lett 95:253703Google Scholar
- Siman L, Carrasco ISS, da Silva JKL, Oliveira MC, Rocha MS, Mesquita ON (2012) Quantitative assessment of the interplay between dna-elasticity and cooperative binding of ligands. Phys Rev Lett 109(24):248103Google Scholar
- Wikipedia (2014) Gelred. http://en.wikipedia.org/wiki/GelRed. Accessed Oct 2014