Circular permutation assay. A) DNA fragments used in the assay. First, the DNA fragment of interest is cloned as a tandem dimer. A variety of restriction enzymes that cut once within the fragment are used to cut the dimer, creating a set of circularly permuted DNA fragments, identical in nucleotide composition and length, but differing in the position of the bend site relative to the fragments' ends. B) Non-denaturing polyacrylamide gel electrophoresis of the circularly permuted DNA fragments. C) Plotting the mobility as a function of the position of cutting. Reproduced from ref. 44 with permission from The Japanese Biochemical Society (©1999).