Transfection of Barley Leaf Protoplasts with a Fluorescently Tagged Fungal Effector for In Planta Localization Studies in Barley

Ana Priscilla Montenegro Alonso; Guus Bakkeren

doi:10.1007/978-1-0716-3159-1_7

Transfection of Barley Leaf Protoplasts with a Fluorescently Tagged Fungal Effector for In Planta Localization Studies in Barley

Methods Mol Biol. 2023:2659:83-93. doi: 10.1007/978-1-0716-3159-1_7.

Authors

Ana Priscilla Montenegro Alonso^{1

2}, Guus Bakkeren³

Affiliations

¹ Department of Botany, University of British Columbia, Vancouver, BC, Canada.
² Saskatoon Research and Development Centre, Agriculture and Agri-Food Canada, Saskatoon, SK, Canada.
³ Summerland Research and Development Centre, Agriculture and Agri-Food Canada, Summerland, BC, Canada. guus.bakkeren@agr.gc.ca.

PMID: 37249887
DOI: 10.1007/978-1-0716-3159-1_7

Abstract

The isolation and transfection of protoplasts from plant leaves have been routinely used for transient expression and functional studies in model plants. However, current approaches to characterize pathogen effector molecules in a cereal host are inefficient and technically challenging. In this chapter, we describe a protocol to isolate and transfect barley mesophyll protoplasts with a fluorescently tagged fungal effector of the barley smut pathogen Ustilago hordei. Tagging of a fungal effector with a fluorescent protein and tracking its localization in cells of its natural host provides insight into its putative in planta localization and helps to narrow down the location of putative host interactors.

Keywords: Barley; Effector; Hordeum vulgare; In planta localization; Leaf protoplast; Microscopy.

MeSH terms

Hordeum* / genetics
Hordeum* / microbiology
Plant Diseases / microbiology
Plant Leaves / genetics
Plant Leaves / microbiology
Protoplasts
Transfection