Melting of Oligodeoxynucleotides with Various Structures

Db, Naritsin; YuL, Lyubchenko

doi:10.1080/07391102.1991.10507847

Cited by 10 publications

(3 citation statements)

References 23 publications

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“…Larger Dumbbells (Naritsin and Lyubchenko, 1990). Recently Naritsin and Lyubchenko (N-L) reported melting studies as a function of "a+] for two DNA dumbbells much larger than the dumbbells of our study.…”

Section: Comparison Of Results With Previous Workmentioning

confidence: 45%

Studies of DNA dumbbells. III. Theoretical analysis of optical melting curves of dumbbells with a 16 base‐pair duplex stem and T_n end loops (n = 2, 3, 4, 6, 8, 10, 14)

1992

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Optical melting curves of seven DNA dumbbells with the 16 base-pair duplex sequence 5'G-C-A-T-A-G-A-T-G-A-G-A-A-T-G-C3' linked on both ends by Tn (n = 2, 3, 4, 6, 8, 10, and 14) loops measured in 30, 70, and 120 mM Na+ are analyzed in terms of the numerically exact statistical thermodynamic model of DNA melting. The construction and characterization of these molecules were described in the previous paper (Amaratunga et al., 1992). As was recently reported for hairpins (T. M. Paner, M. Amaratunga, M. J. Doktycz, and A. S. Benight, 1990, Biopolymers, Vol. 29, pp. 1715-1734) theoretically calculated melting curves were fitted to experimental curves by simultaneously adjusting the parameters representing loop and circle formation to optimize the fits. The systematically determined empirical parameters provide evaluations of the free energies of hairpin loop formation delta Gloop (n) and single-strand circles delta Gcircle (N), as a function of end loop size, n = 2-14, and circle size, N = 32 + 2n. The dependence of these quantities on solvent ionic strength over the range from 30 to 120 mM Na+ was evaluated. An approximately analytical expression for the partition function Q(T) of the dumbbells was formulated that allowed a means for determining the transition enthalpy delta H degrees and entropy delta S degrees for every dumbbell, revealing the dependence of these quantities on loop size. In this multistate approach a manifold of partially melted intermediate microstates are considered and therefore no assumptions regarding the nature of the melting transitions (that they are two-state) are required. The transition thermodynamic parameters were also determined from a van't Hoff analysis of the melting curves. Comparisons between the results of the multistate analysis and the two-state van't Hoff analysis revealed significant differences for the dumbbells with larger end loops, indicating that the melting transitions of the larger looped dumbbells deviate considerably from two-state behavior. Results are then compared with published melting studies of much larger DNA dumbbells (D. B. Naritsin and Y. L. Lyubchenko, 1990, Journal of Biomolecular Structure and Dynamics, Vol. 8, pp. 1-13), of small hairpins (Paner et al., 1990; M. J. Doktycz, T. M. Paner, M. Amaratunga and A. S. Benight, 1990, Biopolymers, Vol. 30, pp. 829-845) and another dumbbell (A. S. Benight, J. M. Schurr, P. F. Flynn, B. R. Reid, and D. E. Wemmer, 1988) Journal of Molecular Biology, Vol. 200, pp. 377-399).(ABSTRACT TRUNCATED AT 400 WORDS)

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Section: Comparison Of Results With Previous Workmentioning

confidence: 45%

Studies of DNA dumbbells. III. Theoretical analysis of optical melting curves of dumbbells with a 16 base‐pair duplex stem and T_n end loops (n = 2, 3, 4, 6, 8, 10, 14)

1992

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“…Hairpin formation, which is not included in the partition function of the model, may contribute to the UV signal at temperatures above the strand dissociation [22]. We examined the stability of the various hairpin structures at different temperatures using the MFOLD server [23].…”

Section: Discussionmentioning

confidence: 99%

Statistical mechanics of base stacking and pairing in DNA melting

2004

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We propose a statistical mechanics model for DNA melting in which base stacking and pairing are explicitly introduced as distinct degrees of freedom. Unlike previous approaches, this model describes thermal denaturation of DNA secondary structure in the whole experimentally accessible temperature range. Base pairing is described through a zipper model, base stacking through an Ising model. We present experimental data on the unstacking transition, obtained exploiting the observation that at moderately low pH this transition is moved down to experimentally accessible temperatures. These measurements confirm that the Ising model approach is indeed a good description of base stacking. On the other hand, comparison with the experiments points to the limitations of the simple zipper model description of base pairing.

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“…On the other hand, all binding sites on a short nucleic acid are close to the molecular ends. This can result in aberrant binding due to structural and electrostatic end-effects [55][56][57] . Longer templates avoid these limitations but contain more non-specific binding sites, migrate more slowly (requiring longer electrophoresis times) and generally give a smaller mobility-shift on protein binding 6 .…”

Section: Strategic Considerations That Are Relevant To Emsamentioning

confidence: 99%

Electrophoretic mobility shift assay (EMSA) for detecting protein–nucleic acid interactions

2007

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The gel electrophoresis mobility shift assay (EMSA) is used to detect protein complexes with nucleic acids. It is the core technology underlying a wide range of qualitative and quantitative analyses for the characterization of interacting systems. In the classical assay, solutions of protein and nucleic acid are combined and the resulting mixtures are subjected to electrophoresis under native conditions through polyacrylamide or agarose gel. After electrophoresis, the distribution of species containing nucleic acid is determined, usually by autoradiography of 32 P-labeled nucleic acid. In general, protein-nucleic acid complexes migrate more slowly than the corresponding free nucleic acid. In this article, we identify the most important factors that determine the stabilities and electrophoretic mobilities of complexes under assay conditions. A representative protocol is provided and commonly used variants are discussed. Expected outcomes are briefly described. References to extensions of the method and a troubleshooting guide are provided.

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Melting of Oligodeoxynucleotides with Various Structures

Cited by 10 publications

References 23 publications

Studies of DNA dumbbells. III. Theoretical analysis of optical melting curves of dumbbells with a 16 base‐pair duplex stem and T_n end loops (n = 2, 3, 4, 6, 8, 10, 14)

Studies of DNA dumbbells. III. Theoretical analysis of optical melting curves of dumbbells with a 16 base‐pair duplex stem and T_n end loops (n = 2, 3, 4, 6, 8, 10, 14)

Statistical mechanics of base stacking and pairing in DNA melting

Electrophoretic mobility shift assay (EMSA) for detecting protein–nucleic acid interactions

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Melting of Oligodeoxynucleotides with Various Structures

Cited by 10 publications

References 23 publications

Studies of DNA dumbbells. III. Theoretical analysis of optical melting curves of dumbbells with a 16 base‐pair duplex stem and Tn end loops (n = 2, 3, 4, 6, 8, 10, 14)

Studies of DNA dumbbells. III. Theoretical analysis of optical melting curves of dumbbells with a 16 base‐pair duplex stem and Tn end loops (n = 2, 3, 4, 6, 8, 10, 14)

Statistical mechanics of base stacking and pairing in DNA melting

Electrophoretic mobility shift assay (EMSA) for detecting protein–nucleic acid interactions

Contact Info

Product

Resources

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Studies of DNA dumbbells. III. Theoretical analysis of optical melting curves of dumbbells with a 16 base‐pair duplex stem and T_n end loops (n = 2, 3, 4, 6, 8, 10, 14)

Studies of DNA dumbbells. III. Theoretical analysis of optical melting curves of dumbbells with a 16 base‐pair duplex stem and T_n end loops (n = 2, 3, 4, 6, 8, 10, 14)