For plasmid manipulations, other site-directed mutagenesis techniques have been supplanted largely by techniques that are highly efficient but relatively simple, easy to use, and commercially available as a kit. An example of these techniques is the Quikchange method, wherein a pair of complementary mutagenic primers are used to amplify the entire plasmid in a thermocycling reaction using a high-fidelity non-strand-displacing DNA polymerase such as "pfu" polymerase. The reaction generates a nicked, circular DNA. The template DNA must be eliminated by enzymatic digestion with a restriction enzyme such as "Dpn"I, which is specific for methylated DNA. All DNA produced from most "Escherichia coli" strains would be methylated; the template plasmid that is biosynthesized in "E. coli" will, therefore, be digested, while the mutated plasmid, which is generated "in vitro" and is therefore unmethylated, would be left undigested. Note that, in these double-strand plasmid mutagenesis methods, while the thermocycling reaction may be used, the DNA need not be exponentially amplified as in a PCR. Instead, the amplification is linear, and it is therefore inaccurate to describe them as a PCR, since there is no chain reaction.