On running a plasmid DNA on 1% Agarose gel, three bands were observed referring to the nicked circular, linear and supercoiled forms. The run difference between these form depend on the plasmid size. Is there a way to determine the difference. Also is there a effective way of determining the apparent size of these bands?
Actually, distances and even position of linear and open circle plasmid DNAs relatively to the supercoiled form depends on several factors. To have a reproducible result, try to use the same lot of agarose. Relative positions of L and OP DNAs depends on the length of DNA and conditions of electrophoresis (buffer, current, temperature, and so on).
Good luck!
Dear Arunkumar
you can try running a supercoiled DNA marker alongside your plasmid preps and the commom linear DNA marker. This way you can estimate the size of the supercoiled plamid DNA.
SD
It depends on the preciseness of the DNA size estimation. You need marker DNA with close linear size. Restriction analysis is more helpful.
Thanks Dr. Drygin and Dr. Datta for your inputs. I ran a supercoiled DNA ladder along with my samples and deduced the sizing of both forms.
Ethidium bromide used in the gel also has a role in the mobilities of different forms of plasmid.
- Badges
- Science topic