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. 2023 Apr;32(1-2):53-66.
doi: 10.1007/s11248-023-00333-1. Epub 2023 Jan 12.

Multiple integrations of a sense transgene, including a tandem inverted repeat confer stable RNA-silencing mediated virus resistance under different abiotic and biotic conditions

Despoina Beris  1 , Aliki Tzima  2 , Fani Gousi  3   2 , Aggeliki Rampou  3 , Venetia Psarra  3 , Ioannis Theologidis  4 , Nikon Vassilakos  3
Affiliations

Multiple integrations of a sense transgene, including a tandem inverted repeat confer stable RNA-silencing mediated virus resistance under different abiotic and biotic conditions

Despoina Beris et al. Transgenic Res. 2023 Apr.

Abstract

In a previous study, tobacco plants, transformed with a sense construct of the 57K domain of the replicase gene of tobacco rattle virus (TRV), provided resistance against genetically distant isolates of the virus. In this work, 57K-specific siRNAs were detected with RT-qPCR solely in the resistant line verifying the RNA-silencing base of the resistance. The integration sites of the transgene into the plant genome were identified with inverse-PCR. Moreover, the resistance against TRV was practically unaffected by low temperature conditions and the presence of heterologous viruses. The mechanism of the resistance was further examined by a gene expression analysis that showed increased transcript levels of genes with a key-role in the RNA silencing pathway and the basal antiviral defence. This work provides a comprehensive characterization of the robust virus resistance obtained by a sense transgene and underlines the usefulness of transgenic plants obtained by such a strategy.

Keywords: I-PCR; Post transcriptional gene silencing (PTGS); Pulsed stem-loop RT; RNAi; Stability of transgenic resistance; Tobravirus.

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