Plum Pox Virus Coat Protein Gene Intron-hairpin-RNA (ihpRNA) Constructs Provide Resistance to Plum Pox Virus in Nicotiana benthamiana and Prunus domestica

Abstract

Constructs with self-complementary sequences separated by an intron produce “hairpin” RNA [intron-hairpin-RNA (ihpRNA)] structures that efficiently elicit posttranscriptional gene silencing (PTGS). In the current study, the authors use this technology to confer resistance to plum pox virus (PPV) in herbaceous and woody perennial plants by silencing the PPV–coat protein (CP) gene. The authors confirmed the high capacity of ihpRNA constructs for inducing RNA silencing in Nicotiana benthamiana Domin., as more than 75% of the transformants displayed PTGS as evaluated by specific small interfering RNA (siRNA) production. The authors demonstrated that ihpRNA constructs provided PPV resistance, and they found a correlation between the length of the PPV sequence introduced in the ihpRNA constructs and the frequency of transgenic-resistant plants. Plants transformed with the full-length sequence produced a higher percentage of resistant lines. The authors further demonstrated for the first time that ihpRNA technology is applicable to a woody perennial species. A transgenic plum (Prunus domestica L.) PPV-CP ihpRNA line showed gene silencing characteristics (hypermethylation of the transgene sequence and specific siRNA production) and resistance to PPV infection 16 months after inoculation.