Abstract
Recent studies have revealed that foliar application of double-stranded RNAs (dsRNAs) or small-interfering RNAs (siRNAs) encoding specific genes of plant pathogens triggered RNA interference (RNAi)-mediated silencing of the gene targets. However, a limited number of reports documented silencing of plant endogenes or transgenes after direct foliar RNA application. This study analyzed the importance of physiological conditions (plant age, time of day, soil moisture, high salinity, heat, and cold stresses) and different dsRNA application means (brush spreading, spraying, infiltration, inoculation, needle injection, and pipetting) for suppression of neomycin phosphotransferase II (NPTII) transgene in Arabidopsis thaliana, as transgenes are more prone to silencing. We observed a higher NPTII suppression when dsRNA was applied at late day period, being most efficient at night, which revealed a diurnal variation in dsRNA treatment efficacy. Exogenous NPTII-dsRNA considerably reduced NPTII expression in 4-week-old plants and only limited it in 2- and 6-week-old plants. In addition, a more discernible NPTII downregulation was detected under low soil moisture conditions. Treatment of adaxial and abaxial leaf surfaces by brushes, spraying, and pipetting showed a higher NPTII suppression, while infiltration and inoculation were less efficient. Thus, appropriate plant age, late time of day, low soil moisture, and optimal dsRNA application modes are important for exogenously induced gene silencing.
Highlights
The discovery of the RNA interference (RNAi) phenomenon has led to the development of powerful genetic engineering tools for crop improvement, disease management, and plant gene functional studies
The main RNAi-based approach relies on constructing transgenic plants expressing double-stranded RNAs (dsRNAs) or hairpin RNA (hpRNA) designed to silence specific plant or plant pathogen genes for regulating plant properties [5,6,14]
Development of new non-invasive technologies based on exogenous RNA treatments with a gene-specific silencing effect that would not permanently modify a plant genome is an important challenge for biotechnology
Summary
RNA interference (RNAi) is a natural gene regulation and antiviral defense mechanism that has been actively explored and exploited for both plant gene functional studies and biotechnological applications in disease control and crop improvement [1,2]. During RNAi, long double-stranded RNAs (dsRNAs) are recognized and converted into small fragments of 20–24-nucleotide (nt)-long RNA duplexes, i.e., small-interfering RNAs (siRNAs) or miRNAs, by a ribonuclease DICER [3,4]. These siRNAs are incorporated into the RNA-induced silencing complex (RISC) to cleave, destabilize, or hinder translation of any homologous mRNAs
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