| Popis: |
The studies in this thesis describe forms of transgenic resistance to plant viruses and how they can be used for studying viral infection cycle. S. tuberosum cv. Saturna expressing the CP gene of Potato mop-top virus (PMTV) was grown in a field infested with the viruliferous vector of PMTV, S. subterranea. The incidence of PMTV-infected tubers was lower in the CP-transgenic potato than in non-transgenic potato. RNA dot-blot analysis revealed that in tubers infected with PMTV, all three RNAs were present. N. benthamiana plants expressing the CP gene of PMTV were inoculated by two different methods i) mechanical inoculation to leaves and ii) growing plants in soil infested with viruliferous S. subterranea. Results showed that the expression of the transgene-derived RNA (or CP) inhibits replication of homologous RNA 2 in transgenic N. benthamiana. Furthermore, the results showed that transgene-mediated resistance to PMTV differs in roots and leaves. Mechanical inoculation with PMTV on CP-transgenic N. benthamiana resulted in symptomless, systemic movement of RNA 1 and RNA 3, but not the CP-encoding RNA (RNA 2). These findings show that PMTV RNA 1 and RNA 3 can infect and move systemically in N. benthamiana without the CP and RNA 2. N. benthamiana transformed with the P1 and VPg cistron, respectively, of Potato virus A (PVA) displayed: i) resistance to PVA infection, ii) susceptibility, or iii) systemic infection followed by recovery from PVA infection of new leaves. Long-distance transport of PVA from lower, infected parts of recovered plants was compromised in the recovered tissue. This result suggests that PVA is moving as ribonucleoprotein complex other than virus particles. N. benthamiana transformed with a polycistronic transgene encoding the CI-NIa-CP cistrons of PVA was susceptible to PVA infection. VPg (the N-proximal part of NIa) is a well-known virulence factor of potyviruses and its possible role in suppression of RNA silencing was studied. PVA VPg was found to increase the severity of disease symptoms when expressed from a Potato virus X vector in N. benthamiana. However, PVA VPg did not show apparent RNA silencing suppression activity. The reason why the polycistronic transgene did not provide resistance could not be resolved. |
