title: Resistance of non-transgenic papaya plants to papaya ringspot virus (PRSV) mediated by intron-containing hairpin dsRNAs expressed in bacteria.
abstract: RNA-mediated virus resistance based on natural antiviral RNA silencing has been exploited as a powerful tool for engineering virus resistance in plants. In this study, a conserved 3'-region (positions 9839-10117, 279 nt) of the capsid protein (CP) gene of papaya ringspot virus (PRSV), designated CP279, was used to generate an intron-containing hairpin RNA (ihpRNA) construct by one-step, zero-background ligation-independent cloning (OZ-LIC). The RNaseIII-deficient Escherichia coli strain M-JM109lacY was identified as the best choice for producing large quantities of specific ihpRNA-CP279. Resistance analyses and ELISA data verified that most papaya plants mechanically co-inoculated with TRIzol-extracted ihpRNA-CP279 and PRSV were resistant to PRSV, and resistance was maintained throughout the test period (>2 months post-inoculation). In contrast, a 1-2 day interval between sequential inoculation of PRSV and ihpRNA-CP279 did not result in complete protection against PRSV infection, but delayed the appearance of viral symptoms by 3 to 4 days. These findings indicate that direct mechanical inoculation of papaya plants with bacterially-expressed ihpRNA-CP279 targeting the PRSV CP gene can interfere with virus infection. This work lays a foundation for developing a non-transgenic approach to control PRSV by directly spraying plants with ihpRNA or crude bacterial extract preparations.
authors.forename: W
authors.initials: W
authors.lastname: Shen
authors.forename: G
authors.initials: G
authors.lastname: Yang
authors.forename: Y
authors.initials: Y
authors.lastname: Chen
authors.forename: P
authors.initials: P
authors.lastname: Yan
authors.forename: D
authors.initials: D
authors.lastname: Tuo
authors.forename: X
authors.initials: X
authors.lastname: Li
authors.forename: P
authors.initials: P
authors.lastname: Zhou
mesh_terms: D029441:Carica / Q000235:genetics / Q000276:immunology* / Q000821:virology; D060467:Disease Resistance; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D007438:Introns; D055029:Inverted Repeat Sequences; D010935:Plant Diseases / Q000276:immunology / Q000821:virology*; D030821:Plants, Genetically Modified / Q000235:genetics / Q000276:immunology* / Q000821:virology; D017800:Potyvirus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D012367:RNA, Viral