Foliar Infiltration of Virus-Derived Small Hairpin RNAs Triggers the RNAi Mechanism Against the Cucumber Mosaic Virus

Overview

Journal Int J Mol Sci

Publisher MDPI

Specialties Biochemistry
Chemistry
Molecular Biology

Date 2022 May 14

PMID 35563329

Authors

Bernardo Villegas-Estrada

Manuel Alejandro Sanchez

Arnubio Valencia-Jimenez

Affiliations

Soon will be listed here.

Abstract

Post-transcriptional gene silencing (PTGS) is an evolutionarily conserved plant defense mechanism against viruses. This paper aimed to evaluate a dsDNA construct (77 bp) as a template for in vitro production of virus-derived artificial small hairpin RNAs (shRNAs) and test for their potential to trigger the RNAi mechanism in plants against CMV after their foliar infiltration. This approach allowed for the production of significant amounts of shRNAs (60-mers) quickly and easily. The gene silencing was confirmed using polymerase chain reaction (PCR), immunological-based assays, and real-time PCR (qPCR). The highest levels of gene silencing were recorded for mRNAs coding for replication protein (ORF1a), the viral suppressor of RNA silencing (ORF2b), and the capsid protein (ORF3b), with 98, 94, and 70% of total transcript silencing, respectively. This protocol provides an alternative to producing significant shRNAs that can effectively trigger the RNAi mechanism against CMV.

Citing Articles

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PMID: 39862243 DOI: 10.1007/s00425-025-04625-0.

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