where ggatcc is BamHI restriction site, ATG is the initial methionine in GOI and ccATGg is NcoI restriction site. Thus, by introducing GOI into pGEM a new restriction site was created.

NcoI recognizes the sequence ccatgg, and DsaI recognizes the sequence ccrygg (r and y stand for any purine and any pyrimidine, respectively). DsaI is less stringent. These two enzymes are not isoschizomers (since Isoschizomers are enzymes that recognize the same pattern).