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Related Experiment Videos

Gateway-compatible vectors for plant functional genomics and proteomics.

Keith W Earley1, Jeremy R Haag, Olga Pontes

  • 1Biology Department, Washington University, 1 Brookings Drive, St Louis, MO 63130, USA. kwearley@artsci.wustl.edu

The Plant Journal : for Cell and Molecular Biology
|January 31, 2006
PubMed
Summary

Gateway cloning offers efficient high-throughput DNA cloning for plants. New pEarleyGate vectors enable easy expression and purification of epitope-tagged proteins in various plant species.

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Area of Science:

  • Molecular Biology
  • Plant Biotechnology

Background:

  • Gateway cloning technology utilizes bacteriophage lambda site-specific recombination for efficient cloning.
  • It enables high-throughput cloning of target sequences into entry and destination vectors for diverse applications.
  • Plant research laboratories widely use Gateway technology for various studies.

Purpose of the Study:

  • To review available Gateway destination vectors for plant research.
  • To introduce pEarleyGate plasmid vectors for Agrobacterium-mediated plant transformation.
  • To demonstrate the utility of pEarleyGate vectors for epitope-tagged protein expression and analysis.

Main Methods:

  • Review of existing Gateway destination vectors.
  • Description of pEarleyGate vectors enabling translational fusion of epitope tags (FLAG, HA, cMyc, AcV5) or tandem affinity purification tags.

Related Experiment Videos

  • Demonstration of pEarleyGate vector utility via immunofluorescence microscopy and affinity capture of epitope-tagged proteins.

    • Main Results:

    • pEarleyGate vectors facilitate expression of epitope-tagged proteins in plants.
    • Epitope tags (FLAG, HA, cMyc, AcV5) show minimal cross-reaction with endogenous plant proteins.
    • Affinity purification and immunolocalization of recombinant proteins are demonstrated.

    Conclusions:

    • Gateway cloning and pEarleyGate vectors provide valuable tools for plant molecular biology research.
    • pEarleyGate vectors simplify the study of protein function, localization, and interactions in plants.
    • The described epitope tags offer broad utility across monocotyledonous and dicotyledonous plants.