Rational design of biosafe crop resistance to a range of nematodes using RNA interference

Abstract

SummaryDouble‐stranded RNA (dsRNA) molecules targeting two genes have been identified that suppress economically important parasitic nematode species of banana. Proteasomal alpha subunit 4 (pas‐4) and Actin‐4 (act‐4) were identified from a survey of sequence databases and cloned sequences for genes conserved across four pests of banana, Radopholus similis, Pratylenchus coffeae, Meloidogyne incognita and Helicotylenchus multicinctus. These four species were targeted with dsRNAs containing exact 21 nucleotide matches to the conserved regions. Potential off‐target effects were limited by comparison with Caenorhabditis, Drosophila, rat, rice and Arabidopsis genomes. In vitro act‐4 dsRNA treatment of R. similis suppressed target gene expression by 2.3‐fold, nematode locomotion by 66 ± 4% and nematode multiplication on carrot discs by 49 ± 5%. The best transgenic carrot hairy root lines expressing act‐4 or pas‐4 dsRNA reduced transcript message abundance of target genes in R. similis by 7.9‐fold and fourfold and nematode multiplication by 94 ± 2% and 69 ± 3%, respectively. The same act‐4 and pas‐4 lines reduced P. coffeae target transcripts by 1.7‐ and twofold and multiplication by 50 ± 6% and 73 ± 8%. Multiplication of M. incognita on the pas‐4 lines was reduced by 97 ± 1% and 99 ± 1% while target transcript abundance was suppressed 4.9‐ and 5.6‐fold. There was no detectable RNAi effect on nontarget nematodes exposed to dsRNAs targeting parasitic nematodes. This work defines a framework for development of a range of nonprotein defences to provide broad resistance to pests and pathogens of crops.

Plant-parasitic nematodes respond to root exudate signals with host-specific gene expression patterns

“…The dsRNAs were designed to be sequence-specific using the assembled genome data to avoid suppression of non-target sequences. A GFP sequence [56] was amplified to provide a control for a non-nematode gene [57]. The DNA fragments were cloned between the XbaI and XhoI sites of the vector L4440 (pPD129.36 [58]).…”

Simultaneous RNAi Knockdown of Three FMRFamide-Like Peptide Genes, Mi-flp1, Mi-flp12, and Mi-flp18 Provides Resistance to Root-Knot Nematode, Meloidogyne incognita

“…The recent introduction of nematicidal chemicals, viz., fluensulfone, fluazaindolizine, tioxazafen, and fluopyrum although have been claimed to be environmentally friendly and well-targeted against RKNs (Kearn et al, 2014;Slomczynska et al, 2014;Faske and Hurd, 2015;Lahm et al, 2017), the adverse effects of chemical pesticides cannot be overruled (Starr et al, 2002). Therefore, there has been a continuing demand for the progression of target-specific, environmentally sound and biodegradable pest management approaches and genetic engineering based techniques have emerged as a precious alternative and corresponding approach in this regard (Fairbairn et al, 2007;Danchin et al, 2013;Roderick et al, 2018;Chaudhary et al, 2019;Hada et al, 2020).…”

Multi-copy alpha-amylase genes are crucial for Ditylenchus destructor to parasitize the plant host

“…As RNAi is a promising strategy for the control of plant-parasitic nematodes, the targets should be designed rationally to avoid off-target effects [ 39 ]. Based on the bioinformatic analyses, the alpha-amylase genes in D .…”