Cleavage of bacteriophage fl DNA by the restriction enzyme of Escherichia coli B.

We studied the cleavage of the replicative-form DNA (RF I) of bacteriophage f1 and its SB mutants by purified restriction endonuclease of E. coli B. The results indicate that: (i) Circular replicative forms are broken once to yield full-length linear molecules (RF III). Such linear molecules are less susceptible than RF I to endonuclease R-B. (ii) The genetic sites (SB sites) that confer on the DNA susceptibility to B-restriction are not the actual sites of cleavage. The number of possible cleavage sites is larger than the number of SB sites. We conclude this because an RF III molecule produced by endonuclease R-B from RF I of a mutant that has only one SB site can be circularized by denaturation and renaturation. (iii) The SB site is not modified when the DNA is cleaved, since an SB site can be used repeatedly by endonuclease R-B; the RF III described in ii can be cleaved by the same enzyme after denaturation and renaturation.