Isolation of Knotted DNA from Coliphage P4

Type II topoisomerases readily remove knots from DNA (1 ). Removal of knots can be assayed by gel electrophoresis (1 ), and this provides a convenient assay for type II topoisomerase activity, even in crude extracts. Such assays have been useful in screening drugs that are suspected to inhibit type II topoisomerases. This chapter describes the isolation of knotted DNA from P4 phage. In the case of wild-type P4, such DNA molecules are double-stranded, hydrogen-bonded, knotted circles of 11,624 nucleotides.