Abstract
Papaya ringspot virus (PRSV) seriously limits papaya (Carica papaya L.) production in tropical and subtropical areas throughout the world. Coat protein (CP)- transgenic papaya lines resistant to PRSV isolates in the sequence-homology-dependent manner have been developed in the U.S.A. and Taiwan. A previous investigation revealed that genetic divergence among Hainan isolates of PRSV has allowed the virus to overcome the CP-mediated transgenic resistance. In this study, we designed a comprehensive RNAi strategy targeting the conserved domain of the PRSV CP gene to develop a broader-spectrum transgenic resistance to the Hainan PRSV isolates. We used an optimized particle-bombardment transformation system to produce RNAi-CP-transgenic papaya lines. Southern blot analysis and Droplet Digital PCR revealed that line 474 contained a single transgene insert. Challenging this line with different viruses (PRSV I, II and III subgroup) under greenhouse conditions validated the transgenic resistance of line 474 to the Hainan isolates. Northern blot analysis detected the siRNAs products in virus-free transgenic papaya tissue culture seedlings. The siRNAs also accumulated in PRSV infected transgenic papaya lines. Our results indicated that this transgenic papaya line has a useful application against PRSV in the major growing area of Hainan, China.
Highlights
Papaya (Carica papaya L.) is a widely cultivated fruit crop in the tropics and sub-tropics
The genetic variability among Papaya ringspot virus (PRSV) isolates in Hainan was investigated in a previous study[16]
Breakdown of transgenic PRSV resistance, which depends on sequence homology between the transgene and attacking virus strain, is a major concern facing papaya cultivation, since genetically distinct strains of PRSV have been identified throughout the world[18]
Summary
Papaya (Carica papaya L.) is a widely cultivated fruit crop in the tropics and sub-tropics. Genome sequence analysis revealed that the papaya has significantly fewer disease resistance genes than that of Arabidopsis. Transgenic papaya cultivars have varying levels of resistance against PRSV isolates from other geographical regions. Resistance levels depend on the variability among CP genes of the isolates[8,14,15]. The high levels of genetic divergence in PRSV isolates from Hainan is likely to be the cause of the failure of transgenic papaya lines that targets specific viral CP genes[16]. Transgenic papaya has been proven to have effective resistance to PRSV isolates from Hawaii, Taiwan, and other. We use the RNAi strategy to construct a transgene that targets the conserved region of the PRSV CP genes and confers broad-spectrum resistance to the diverse Hainan PRSV isolates
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