title: Improving the efficiency of transposon mutagenesis in Salmonella enteritidis by overcoming host-restriction barriers.
abstract: Transposon mutagenesis using transposome complex is a powerful method for functional genomics analysis in diverse bacteria by creating a large number of random mutants to prepare a genome-saturating mutant library. However, strong host restriction barriers can lead to limitations with species- or strain-specific restriction-modification systems. The purpose of this study was to enhance the transposon mutagenesis efficiency of Salmonella Enteritidis to generate a larger number of random insertion mutants. Host-adapted Tn5 DNA was used to form a transposome complex, and this simple approach significantly and consistently improved the efficiency of transposon mutagenesis, resulting in a 46-fold increase in the efficiency as compared to non-adapted transposon DNA fragments. Random nature of Tn5 insertions was confirmed by high-throughput sequencing of the Tn5-junction sequences. The result based on S. Enteritidis in this study should find broad applications in preparing a comprehensive mutant library of other species using transposome complex.
authors: Cell and Molecular Biology Program, University of Arkansas, Fayetteville, AR, 72701, USA, dturki@gmail.com.
authors: Turki M
authors: TM
authors: Dawoud
authors: Tieshan
authors: T
authors: Jiang
authors: Rabindra K
authors: RK
authors: Mandal
authors: Steven C
authors: SC
authors: Ricke
authors: Young Min
authors: YM
authors: Kwon
mesh_terms: D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics; D016254:Mutagenesis, Insertional / Q000379:methods*; D012477:Salmonella enteritidis / Q000235:genetics*; D017422:Sequence Analysis, DNA
chemical_list: D004251:DNA Transposable Elements; D004269:DNA, Bacterial