FIG. 5.

Alteration of nucleotides inside the 28-bp IES does not prevent excision. Four nucleotides inside the 28-bp IES were altered to create a unique SacI restriction site in pIES-SacI (B). After injection of pIES-SacI into mating cells and isolation of DNA from drug-resistant transformants, amplified PCR products were separated on a 4% agarose gel (A). (A) Agarose gel stained with ethidium bromide (top) and Southern hybridization of the blotted gel using the 28-bp IES junction specific oligonucleotide (see Materials and Methods) (bottom). Lane 1 contains product from pIES-SacI.