Figure 5

Transfer of the esp PAI from E. faecium E1162Δ esp to E. faecium BM4105RF. Panel (A): representative ethidium bromide stained PFGE gel of SmaI-digested chromosomal DNA of the donor strain (E1162Δesp; lane 1), the recipient strain (BM4105RF; lane 2) and a transconjugant (lane 3). The gel band that has shifted in the recipient strain due to the insertion of the esp PAI is indicated by the white arrow. Panel (B): Southern blots of the PFGE gel hybridized with a probe for esp and a probe covering the insertion site of the esp PAI. Panel (C): PCR analysis of the esp PAI insertion site in the donor strain (E1162Δesp), the recipient strain (BM4105RF) and a transconjugant (TC). PCRs were performed with primers covering the esp PAI insertion site (reactions A), covering the 5' end of the esp PAI and the 5' flanking end of the insertion site (reactions B), and covering the 3' end of the esp PAI and the 3' flanking end of the insertion site (reactions C). The marker (M) is Invitrogen's 1 Kb Plus DNA Ladder. Note that, because of the size of the esp PAI, no products can be obtained in PCRs with primers on the 5' and 3' flanking regions of the esp PAI integration site in esp⁺ strains.