A target bacterial strain of interest for use in Red-based recombineering may already encode resistance to antibiotic markers used with current Red recombination tools, such that the resistance cannot be removed. Such cases include those where markers are needed to maintain an unstable genetic element co-resident in the strain or those where the genetic source of resistance is not known. We report the availability of PCR templates with FRT-flanked mutagenesis cassettes and plasmids encoding Red recombination functions that contain marker combinations not currently available on widely disseminated lambda Red molecular reagents. The functionality of these convenient alternative tools is demonstrated.