FIG. 1.

Map of the bfp gene cluster illustrating construction of bfpF mutants. (A) The entire bfp gene cluster is shown. Arrows represent bfp genes. Selected restriction enzyme sites shown in bfpF were used for cloning. (B) The bfpEFP genes are magnified to illustrate the construction of pRPAF1. A 72-bp KpnI site was deleted from a subcloned fragment containing bfpF as shown in the line below the map. The SacI-DraIII fragment from this construct was then used to replace the SacI-DraIII fragment of pKDS135 to generate pRPAF1. (C) The bfpEFP genes are magnified to illustrate the construction of strain UMD916. A nonpolar kanamycin resistance cassette containing the aphA3 gene was inserted via its flanking SmaI sites into the BbrPI site in a subcloned fragment containing bfpF. B/S and S/B represent the junctions of ligated fragments that had BbrPI and SmaI sites. A fragment containing this insert was cloned into positive-selection suicide vector pCVD442 for allelic exchange with the wild-type strain.