Fig. 2.
BbvCI reactions. (a) Shown as ABC is a 301-bp DNA with two BbvCI sites in inverted orientation 75 bp apart. The sites, 1 and 2, have the scissile bonds in the CC sequence at positions 101 and 173, respectively. The initial reaction of BbvCI on this DNA has three possible outcomes: cleavage at site 1 alone to give fragments A (in black) and BC (in blue); at 2 alone for fragments AB (in red) and C (in green); processive cutting at both 1 and 2 to give fragments A, B (in purple), and C. (b and c) BbvCI endonuclease (0.3 nM) was added to the above DNA (32P-labeled, 30 nM) in Reaction Buffer at 37°C. Samples were taken at the indicated times and subjected to electrophoresis through polyacrylamide. b shows a phosphorimager record of the gel, annotated on the right with the mobilities (and sizes) of all of the DNA species from the reaction. For c, the concentrations of the following fragments were evaluated from the record: A, black diamonds; BC, blue circles; AB, red triangles; C, green squares. The dotted lines through each data set are the optimal fits to linear slopes to give, respectively, vA, vBC, vAB, and vC.