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Generation of Conditional Mutants in Borrelia burgdorferi

  • Jacob I. Latham
  • Jon S. Blevins
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1690)

Abstract

Mutational studies aimed at characterizing the function(s) of bacterial genes required for growth or viability are constrained by the inability to generate deletion strains lacking the gene of interest. To circumvent this limitation, it is possible to generate conditional mutants in which a copy of the gene of interest is introduced into the bacteria to compensate for the loss of the native allele. Expression of the non-native copy of the target gene is typically under control of an inducible promoter, which allows for controllable and regulated production of the gene of interest. Conditional mutants are also broadly useful for phenotypic analyses of genes that require a tightly regulated and artificially inducible copy of the target gene. Herein, we describe the methods used to generate and confirm conditional mutant clones in Borrelia burgdorferi utilizing the Borrelia-adapted lac operator/repressor system.

Key words

Borrelia Lyme disease Inducible expression Conditional mutants Genetic manipulation lac repressor LacI Essential genes 

Notes

Acknowledgments

This work was supported by funding to J.S.B. through the Arkansas Biosciences Institute (major research component of the Arkansas Tobacco Settlement Proceeds Act of 2000), NIH/NIAID R01-AI087678, NIH/NIAID R21-AI 119532, as well as support through the Translational Research Institute (UL1-TR000039; NIH National Center for Research Resources and National Center for Advancing Translational Sciences) and the UAMS Center for Microbial Pathogenesis and Host Inflammatory Responses (P20-GM103625).

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Copyright information

© Springer Science+Business Media LLC 2018

Authors and Affiliations

  1. 1.Department of Microbiology and ImmunologyUniversity of Arkansas for Medical SciencesLittle RockUSA

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