The linear form is a result of a cleavage on both DNA strands caused by restriction endonucleases.Īn open circle (OC) dimer is an oligomeric form of a plasmid. It also has less supercoiling than the covalently closed circular form.

This structure is a relaxed and less compact form of plasmid. UV irradiation or nucleases can cause this single-strand break. Undigested plasmid DNA are usually supercoiled.Īn open circular form is caused by the nicking (cleavage) of one DNA strand. Plasmid DNA isolated from bacterial hosts are usually present in this covalently closed circular form. Intact supercoiled plasmids have compact double-stranded DNA twisted around itself.

The covalently closed circular monomer is a negatively charged, supercoiled plasmid. Supercoiled DNA are more difficult to trap due to the small size of the twisted DNA.Ĥ Common Forms of Plasmid DNA Covalently Closed Circle(CCC) Monomer So, large circular molecules have a greater chance to get trapped than smaller DNA forms. The electrophoretic trapping is a balance between the electrophoretic force (pulling the circular plasmid DNA against the trap) and diffusion (allowing the circular plasmid DNA to escape a trap). The gel electrophoresis conditions, including the presence of ethidium bromide (or alternative), gel concentrations, electric field strength, temperature, and ionic strength of the electrophoresis buffer, can affect the mobility of plasmid DNA. The travel distance of DNA molecules within an agarose gel is proportional to the log of its molecular weight. Because of the negatively charged phosphate backbone, DNA holds a slight negative charge that allows it to migrate to the positively charged anode. Under a powerful microscope, a gel will look porous, but to the naked eye, it looks like a smooth, opaque gelatin in the shape of a square with wells near one end of the surface.Ī well is a hollow pocket in the gel where the DNA is loaded. Let’s look at how DNA electrophoresis in an agarose gel works. Smaller DNA fragments can move quickly through the pores, while larger fragments get caught and therefore travel slowly. ĭNA separation occurs due to the mesh-like nature of the agarose gel. This network consists of pores with molecular filtering properties.Ĭonceptual rendering of agarose gel at a microscopic level. During polymerization, agarose polymers link non-covalently and form a network of bundles. Tips To Identify The Bands In Your Agarose GelĪgarose, produced from seaweed, is a polysaccharide. How Does Circular Plasmid DNA Run During Gel Electrophoresis? In this article, we will review the different forms of plasmid DNA and offer some useful tips to interpret your gel. However, when you look at your gel, you may see multiple bands in a given lane and wonder which one you should cut. When you use gel electrophoresis to help you with molecular cloning, you will also need to be able to interpret and analyze the results of your gel.įor example, you may need to excise your digested plasmid DNA from agarose. Gel electrophoresis is a molecular biology method used to analyze and separate DNA fragments based on their size.

Comparison of dna sequences in table ii