Transplastomic Tomato Plants Expressing Insect-Specific Double-Stranded RNAs: A Protocol Based on Biolistic Transformation

Emine Kaplanoglu; Igor Kolotilin; Rima Menassa; Cam Donly

doi:10.1007/978-1-0716-1633-8_18

Transplastomic Tomato Plants Expressing Insect-Specific Double-Stranded RNAs: A Protocol Based on Biolistic Transformation

Methods Mol Biol. 2022:2360:235-252. doi: 10.1007/978-1-0716-1633-8_18.

Authors

Emine Kaplanoglu¹, Igor Kolotilin², Rima Menassa^{1

3}, Cam Donly^{4

5}

Affiliations

¹ London Research and Development Centre, Agriculture and Agri-Food Canada, London, ON, Canada.
² Solar Grants Biotechnology Inc, London, ON, Canada.
³ Department of Biology, University of Western Ontario, London, ON, Canada.
⁴ London Research and Development Centre, Agriculture and Agri-Food Canada, London, ON, Canada. cam.donly@canada.ca.
⁵ Department of Biology, University of Western Ontario, London, ON, Canada. cam.donly@canada.ca.

PMID: 34495519
DOI: 10.1007/978-1-0716-1633-8_18

Abstract

Expressing insecticidal double-stranded RNA (dsRNA) molecules in plant plastids is a novel approach for in planta production of dsRNA that has enormous potential for developing improved plant-mediated RNA interference (RNAi) strategies for insect pest control. In this chapter, we describe the design of a transformation vector containing an expression cassette which can be used to stably transform plastids of tomato plants for production and accumulation of dsRNA . Such dsRNA can trigger the mechanisms of RNAi in pest insects and selectively suppress the expression of target genes, resulting in lethality. We also describe a protocol for detection of full-length dsRNA molecules in plastids using an RT-PCR-based method.

Keywords: Biolistic delivery; Double-stranded RNA detection; Insect pest management; Plastid transformation; RNA interference; RT-PCR.

Publication types

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

MeSH terms

Animals
Biolistics*
Insecta / genetics
RNA Interference
RNA, Double-Stranded / genetics
RNA, Plant / genetics
Solanum lycopersicum* / genetics

Substances

RNA, Double-Stranded
RNA, Plant