Abstract
Markerless chromosomal in-frame deletion mutants are often used to avoid polar effects, conserve antibiotic resistance markers or make multiple mutations. This protocol capitalizes on the sacB gene (lethal in the presence of sucrose), for selection against a single cross-over event, on a suicide vector (contains no Rhodobacter capsulatus compatible origin of replication). The bacterial strains, media and suicide vectors in this protocol are specific for R. capsulatus, but the concepts can be applied to other species.
Materials and Reagents
Equipment
Procedure
Recipes
Acknowledgments
The development of this protocol was funded by a grant to J.T.B. from the Canadian Institutes of Health Research. This protocol was adapted from protocols developed by Beatty and Gest (1981), and Penfold and Pemberton (1992).
References
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