London Research and Development Centre, Agriculture and Agri-Food Canada, London, ON, Canada.
Solar Grants Biotechnology Inc, London, ON, Canada.
Methods Mol Biol. 2022;2360:235-252. doi: 10.1007/978-1-0716-1633-8_18.
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.
在植物质体中表达杀虫双链 RNA (dsRNA) 分子是一种在植物体内生产 dsRNA 的新方法，对于开发改进的基于植物的 RNA 干扰 (RNAi) 策略来控制昆虫害虫具有巨大的潜力。在本章中，我们描述了一个包含表达盒的转化载体的设计，该载体可用于稳定转化番茄植物的质体，以生产和积累 dsRNA。这种 dsRNA 可以触发害虫体内的 RNAi 机制，并选择性地抑制靶基因的表达，导致其死亡。我们还描述了一种使用基于 RT-PCR 的方法检测质体中全长 dsRNA 分子的方案。
