Two-dimensional agarose gel electrophoresis of DNA topoisomers.
The electrophoretic velocity of a duplex DNA ring is mainly determined by its overall shape. Consequently, DNA topoisomers of opposite supercoiling handedness can have identical gel velocity, and topoisomers highly supercoiled cannot be separated beyond some point. These problems are overcome by two-dimensional agarose gel electrophoresis, which involves two successive electrophoresis steps in one gel slab. The first and second electrophoresis steps are conducted in orthogonal directions with different concentrations of DNA intercalating agents. These compounds alter the overall shape of the DNA and, thereby, change the relative mobility of individual DNA topoisomers.
Roca
Joaquim
J
IBMB-CSIC, Barcelona, Spain.
D004247:DNA / Q000737:chemistry*; D026942:DNA Topoisomerases / Q000737:chemistry; D004278:DNA, Superhelical / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional / Q000295:instrumentation / Q000379:methods*; D007364:Intercalating Agents / Q000737:chemistry; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000737:chemistry / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics
D004278:DNA, Superhelical; D007364:Intercalating Agents; D004247:DNA; D026942:DNA Topoisomerases