Recombineering and stable integration of the Pseudomonas syringae pv. syringae 61 hrp/hrc cluster into the genome of the soil bacterium Pseudomonas fluorescens Pf0-1

Authors

  • William J. Thomas,

    1. Department of Botany and Plant Pathology, Oregon State University, Corvallis, OR 97331, USA
    2. Molecular and Cellular Biology Program, Oregon State University, Corvallis, OR 97331, USA
    Search for more papers by this author
    • These authors contributed equally to this work.

  • Caitlin A. Thireault,

    1. Department of Botany and Plant Pathology, Oregon State University, Corvallis, OR 97331, USA
    Search for more papers by this author
    • These authors contributed equally to this work.

  • Jeffrey A. Kimbrel,

    1. Department of Botany and Plant Pathology, Oregon State University, Corvallis, OR 97331, USA
    2. Molecular and Cellular Biology Program, Oregon State University, Corvallis, OR 97331, USA
    Search for more papers by this author
  • Jeff H. Chang

    Corresponding author
    1. Department of Botany and Plant Pathology, Oregon State University, Corvallis, OR 97331, USA
    2. Molecular and Cellular Biology Program, Oregon State University, Corvallis, OR 97331, USA
    3. Center for Genome Research and Biocomputing, Oregon State University, Corvallis, OR 97331, USA
    Search for more papers by this author

(fax 541 737 3573; e-mail changj@science.oregonstate.edu).

Summary

Many Gram-negative bacteria use a type III secretion system (T3SS) to establish associations with their hosts. The T3SS is a conduit for direct injection of type-III effector proteins into host cells, where they manipulate the host for the benefit of the infecting bacterium. For plant-associated pathogens, the variations in number and amino acid sequences of type-III effectors, as well as their functional redundancy, make studying type-III effectors challenging. To mitigate this challenge, we developed a stable delivery system for individual or defined sets of type-III effectors into plant cells. We used recombineering and Tn5-mediated transposition to clone and stably integrate, respectively, the complete hrp/hrc region from Pseudomonas syringae pv. syringae 61 into the genome of the soil bacterium Pseudomonas fluorescens Pf0-1. We describe our development of Effector-to-Host Analyzer (EtHAn), and demonstrate its utility for studying effectors for their in planta functions.

Ancillary