Vacuole-Targeted Proteins: Ins and Outs of Subcellular Localization Studies

Methods Mol Biol. 2018:1789:33-54. doi: 10.1007/978-1-4939-7856-4_4.

Abstract

Accurate and efficient demonstrations of protein localizations to the vacuole or tonoplast remain strict prerequisites to decipher the role of vacuoles in the whole plant cell biology and notably in defence processes. In this chapter, we describe a reliable procedure of protein subcellular localization study through transient transformations of Catharanthus roseus or onion cells and expression of fusions with fluorescent proteins allowing minimizing artefacts of targeting.

Keywords: Allium cepa; Biolistic; Catharanthus roseus; GFP; Onion peel; Protein localization; Tonoplast; Transient transformation; Vacuolar targeting; Vacuole; YFP.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Proteins / analysis*
  • Bacterial Proteins / genetics
  • Catharanthus / cytology*
  • Catharanthus / genetics
  • Genetic Vectors / genetics
  • Green Fluorescent Proteins / analysis*
  • Green Fluorescent Proteins / genetics
  • Luminescent Proteins / analysis*
  • Luminescent Proteins / genetics
  • Microscopy, Fluorescence / methods
  • Onions / cytology*
  • Onions / genetics
  • Plant Proteins / analysis*
  • Plant Proteins / genetics
  • Protein Transport
  • Recombinant Fusion Proteins / analysis
  • Recombinant Fusion Proteins / genetics
  • Transformation, Genetic
  • Vacuoles / chemistry
  • Vacuoles / genetics
  • Vacuoles / ultrastructure*

Substances

  • Bacterial Proteins
  • Luminescent Proteins
  • Plant Proteins
  • Recombinant Fusion Proteins
  • yellow fluorescent protein, Bacteria
  • Green Fluorescent Proteins