Quantification of DNA by agarose gel electrophoresis and analysis of the topoisomers of plasmid and M13 DNA following treatment with a restriction endonuclease or DNA topoisomerase I

Abstract

A two-session laboratory exercise for advanced undergraduate students in biochemistry and molecular biology is described. The first session introduces students to DNA quantification by ultraviolet absorbance and agarose gel electrophoresis followed by ethidium bromide staining. The second session involves treatment of various topological forms of DNA with a restriction endonuclease and with a DNA topoisomerase. This session introduces students to the concept of DNA topoisomers, to the properties of different forms of DNA, and to the activity of restriction endonucleases and topoisomerases toward these forms. The exercise also involves measuring the size of linear duplex fragments of DNA by comparison of mobility with a ladder of double stranded DNA of known sizes.