Fast-Forward Identification of Highly Effective Artificial Small RNAs Against DifferentTomato spotted wilt virusIsolates

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Abstract
Artificial small RNAs (sRNAs), including artificial microRNAs (amiRNAs) and synthetic trans-acting small interfering RNAs (syn-tasiRNAs), are used to silence viral RNAs and confer antiviral resistance in plants. Here, the combined use of recent high-throughput methods for generating artificial sRNA constructs and the Tomato spotted wilt virus (TSWV)–Nicotiana benthamiana pathosystem allowed for the simple and rapid identification of amiRNAs with high anti-TSWV activity. A comparative analysis between the most effective amiRNA construct and a syn-tasiRNA construct including the four most effective amiRNA sequences showed that both were highly effective against two different TSWV isolates. These results highlight the usefulness of this high-throughput methodology for the fast-forward identification of artificial sRNAs with high antiviral activity prior to time-consuming generation of stably transformed plants.
Funding Information
  • Ministerio de Economía y Competitividad (BIO2014-54269-R)
  • Ministerio de Ciencia, Innovación y Universidades (BIO2017-83184-R)
  • Instituto Nacional de Investigaciones Agrarias (RTA2013-00047-C02-02)
  • Marie Sklodowska Curie Individual Fellowship, European Union Horizon 2020 (655841)

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