Comparative Analysis of RNAi-Based Methods to Down-Regulate Expression of Two Genes Expressed at Different Levels in Myzus persicae.

Michaël Mulot,Véronique Brault,Nicole Bochet,Sylvaine Boissinot,Maryam Rastegar,Baptiste Monsion,Gabriel Clavijo,David Halter,Monique Erdinger

doi:10.3390/v8110316

Abstract

With the increasing availability of aphid genomic data, it is necessary to develop robust functional validation methods to evaluate the role of specific aphid genes. This work represents the first study in which five different techniques, all based on RNA interference and on oral acquisition of double-stranded RNA (dsRNA), were developed to silence two genes, ALY and Eph, potentially involved in polerovirus transmission by aphids. Efficient silencing of only Eph transcripts, which are less abundant than those of ALY, could be achieved by feeding aphids on transgenic Arabidopsis thaliana expressing an RNA hairpin targeting Eph, on Nicotiana benthamiana infected with a Tobacco rattle virus (TRV)-Eph recombinant virus, or on in vitro-synthesized Eph-targeting dsRNA. These experiments showed that the silencing efficiency may differ greatly between genes and that aphid gut cells seem to be preferentially affected by the silencing mechanism after oral acquisition of dsRNA. In addition, the use of plants infected with recombinant TRV proved to be a promising technique to silence aphid genes as it does not require plant transformation. This work highlights the need to pursue development of innovative strategies to reproducibly achieve reduction of expression of aphid genes.

Highlights

Aphids are small sap-sucking insects belonging to the Hemiptera order
In order to evaluate the efficacy of different RNA interference (RNAi)-based silencing techniques in aphids, we selected two genes from M. persicae, Ephrin receptor (Eph) and ALY, that are potentially involved in polerovirus transmission
ALY transcripts accumulated at a higher level compared to Eph mRNA in non-viruliferous M. persicae, which provided the opportunity to compare the efficacy of the silencing techniques with two genes that have different baseline levels of transcript abundance

Summary

Introduction

Aphids are small sap-sucking insects belonging to the Hemiptera order. They display important demographic potential and adapt to changes in environmental conditions, causing significant damage to crops by direct feeding. They are vectors of numerous deleterious plant viruses, and are considered as one of the main animal pests for agriculture [1,2,3]. Mainly pesticide use, have been deployed in the field to reduce aphid populations, but the development of insecticide resistance has been an increasing problem for agriculture [3,4,5].

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Conclusion