In Site directed mutagenesis using PCR, after a cycle we obtain a hybrid molecule with one parental strand and other newly synthesized unmethylated strand. This is followed by Dpn1 digestion. Does this digestion just cut the parental strand leaving behind single strand of newly synthesized product? Does this means that Dpn1 can use ds methylated DNA( both strands methylated) as well as ds hybrid DNA (with one methylated strand and other unmethylated strand) as substrate?
According to the NEB website FAQ, yes, it can, but less efficiently.
"DpnI cleaves hemi-methylated dam sites 60X more slowly than fully methylated."