Tn-seq

Tn-seq identifies essential genes within specific environments by mapping genome-wide transposon insertions through high-throughput sequencing of transposon-genome junctions.

Key Features:

• Transposon libraries: Generation of genome-wide transposon insertion libraries to inactivate nonessential genes.
• Random insertion mapping: Integration of transposons into random genomic locations to assay gene essentiality.
• Sequencing platform: Sequencing of transposon-genome junctions using Illumina high-throughput sequencing technologies.
• Computational processing: Scripts developed in the Whiteley lab automate parsing, mapping, and tallying of Tn-seq reads.
• Differential abundance analysis: Identification of differentially abundant transposon insertion mutants to determine essential genes.
• Cross-strain comparison: Application to comprehensive libraries across multiple strains to compare gene essentiality in different genetic backgrounds.

Scientific Applications:

• Staphylococcus aureus gene essentiality: Identification of essential genes in Staphylococcus aureus strains.
• Host-environment essentiality: Determination of genes required for bacterial survival in specific host environments.
• Animal infection models: Use in animal models of infection, for example murine models of osteomyelitis, to identify genes necessary for in vivo survival.
• Pathogenesis mechanisms: Pinpointing genes indispensable for bacterial pathogenesis within host tissues.

Methodology:

Sequencing of transposon-genome junctions (Illumina) produces datasets that are parsed, mapped, and tallied by scripts developed in the Whiteley lab, followed by analysis to identify differentially abundant transposon insertion mutants.