1a. Objectives (from AD-416):
1) Develop qPCR probes for the different Dekopon sub-isolates and quantify each in single and mixed infections to assess interference in accumulation/replication of each sub-isolate. 2) Examine P20, P23, and CP (suppressors of gene silencing) for each sub-isolate for genetic variation and associate with symptom expression. 3) Characterize siRNA profiles in mixed and single infected plants.
1b. Approach (from AD-416):
Aphid transmission, reverse transcription (RT) polymerase chain reaction (PCR), real time PCR (qPCR), cloning, sequencing, western blots, northern blots, hybridizations.
3. Progress Report:
Results from this study are in support of objective 3A of the parent project. Viral cross protection is a natural plant defense system where a mild virus strain prevents subsequent infection of a second, more severe strain of the same virus. The mechanism of cross-protection is unknown, however, one possible mechanism may be RNA interference. During FY12, cross-protection of severe strains of Citrus Tristeza Virus was examined in citrus plants infected with a mixture of two strains (one mild and the other severe). Since RNA interference involves inactivation of the virus by cleaving viral RNA into short segments (21-28 nucleotides), a study was conducted to characterize small viral RNAs isolated from plants in cross-protection tests. Infected, but asymptomatic plants (protected) contained predominately small viral RNAs from the severe strain. In contrast, infected plants with severe symptoms (non-protected) had prominent small viral RNAs from the mild strain. The data supports the hypothesis that cross-protection occurs when the plant host RNA interference system targets the RNA from the severe strain viral for degradation.