Specialized transduction designed for precise high-throughput unmarked deletions in Mycobacterium tuberculosis.

ABSTRACT:

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Specialized transduction has proven to be useful for generating deletion mutants in most mycobacteria, including virulent Mycobacterium tuberculosis. We have improved this system by developing (i) a single-step strategy for the construction of allelic exchange substrates (AES), (ii) a temperature-sensitive shuttle phasmid with a greater cloning capacity than phAE87, and (iii) bacteriophage-mediated transient expression of site-specific recombinase to precisely excise antibiotic markers. The methods ameliorate rate-limiting steps in strain construction in these difficult-to-manipulate bacteria. The new methods for strain construction were demonstrated to generalize to all classes of genes and chromosomal loci by generating more than 100 targeted single- or multiple-deleti

SUBMITTER: Jain P 

PROVIDER: S-EPMC4049104 | biostudies-literature | 2014 Jun

REPOSITORIES: biostudies-literature