Education
with Oleg Krichevsky
with Oleg Krichevsky
Show abstract
The structure of circular DNA (plasmids) is determined by its superhelicity, a topological
quality which can be thought of as the excess, or defficiency, of whole twist turns of the double
helix around its axis. Since supercoiled plasmids are a few hundred nanometers in size, their
structural details cannot be observed optically. While there have been studies to characterize
the structure of supercoiled plasmids, none have attempted to probe the structure in a liquid
solution, resembling physiological conditions.
We present an in-vitro study of the structure of supercoiled plasmids in solution: Plasmids
are labeled uniformly and covalently with minimal structural interference. A combination
of scanning
Fuorescent correlation spectroscopy (sFCS) and time-gated stimulated emission
depletion microscopy (gSTED) are utilized to measure the static structure factor of plasmid
topoisomers, and its dependence on superhelicity. By analyzing the results and comparing
with a DNA simulation, we shed more light on the actual structure of supercoiled plasmids
- support for a branched plectonemic structure is given, and a qualitative resemblance is
observed between the measured and simulated gyration radii and structure factors of both
relaxed and supercoiled plasmids.