Related Experiment Videos
Atomic force microscopy studies of intercalation-induced changes in plasmid DNA tertiary structure
L H Pope1, M C Davies, C A Laughton
1Laboratory of Biophysics and Surface Analysis, School of Pharmaceutical Sciences, The University of Nottingham, University Park, Nottingham NG7 2RD, U.K.
Journal of Microscopy
|July 8, 2000
Summary
Atomic force microscopy revealed that plasmid DNA undergoes structural transitions. Increasing ethidium bromide concentration shifted DNA from relaxed states to toroidal and then plectonemic supercoiling.
Area of Science:
- Structural biology
- Biophysics
- Molecular biology
Background:
- Plasmid DNA exists in various supercoiled tertiary structures.
- Superhelical stress influences DNA conformation and function.
- Ethidium bromide is a known intercalator that alters DNA topology.
Purpose of the Study:
- To investigate structural transitions in plasmid DNA tertiary structure.
- To monitor conformational changes induced by ethidium bromide intercalation.
- To characterize the supercoiling states adopted by plasmid DNA.
Main Methods:
- Atomic force microscopy (AFM) was employed to visualize individual DNA molecules.
- Topographic images of DNA-ethidium bromide complexes were recorded.
- Varying ethidium bromide to base pair ratios were used to induce superhelical stress.
Main Results:
- Significant tertiary structure changes were observed with increasing ethidium bromide concentration.
- A transition from relaxed DNA to toroidal supercoiling was identified.
- Further intercalation led to the formation of plectonemic supercoiling, eventually dominating the structure.
Conclusions:
- Plasmid DNA exhibits distinct structural transitions in response to ethidium bromide-induced superhelical stress.
- The observed toroidal supercoiling is an unusual intermediate state, contrasting with previously established plectonemic dominance.
- AFM provides high-resolution insights into DNA structural dynamics and topological transitions.