Isolation and Transfection of Plant Mesophyll Protoplasts for Virology Research

Zhaoji Dai; Aiming Wang

doi:10.1007/978-1-0716-1835-6_5

Isolation and Transfection of Plant Mesophyll Protoplasts for Virology Research

Methods Mol Biol. 2022:2400:43-53. doi: 10.1007/978-1-0716-1835-6_5.

Authors

Zhaoji Dai^{1

2}, Aiming Wang³

Affiliations

¹ London Research and Development Centre, Agriculture and Agri-Food Canada, London, ON, Canada.
² Department of Biology, University of Western Ontario, London, ON, Canada.
³ London Research and Development Centre, Agriculture and Agri-Food Canada, London, ON, Canada. aiming.wang@canada.ca.

PMID: 34905189
DOI: 10.1007/978-1-0716-1835-6_5

Abstract

Protoplasts are the naked plant cells lacking the rigid cell wall and have been broadly utilized as an excellent tool to study the molecular virus-plant interactions, particularly at the early stages of the infection process, such as virion disassembly, viral genome translation, intracellular trafficking, and virus replication. Compared to the use of whole plants, the protoplast system has several major advantages in plant virology research, including homogeneous cell populations, high percentage of infected cells, synchronous infection, effects free from other cells/tissues, and ease of extraction of the viral RNA. This chapter describes a simple, streamlined, and efficient protocol for isolation and purification of mesophyll protoplasts from the model plants Arabidopsis thaliana and Nicotiana benthamiana, and subsequent transfection of the isolated protoplasts with a potyvirus infectious clone.

Keywords: Arabidopsis thaliana; Mesophyll protoplast; Nicotiana benthamiana; Plant virus infection; Potyvirus; Transfection.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Arabidopsis / genetics
Cell Wall
Nicotiana
Plant Viruses
Protoplasts*
Transfection