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 by high throughput sequencing.
1b. Approach (from AD-416)
3. Progress Report
This Specific Cooperative Agreement was established in support of Objective 1B. The goal is to identify genes that regulate disease expression of Citrus tristeza virus (CTV). This research contributes to development of control measures for CTV and advances knowledge on the role of gene silencing in mild strain cross protection of CTV against expression or co-infection by virulent strains. Evaluation of micro RNA profiles in cross protected vs. non cross-protected sour orange plants identified putative host genes targeted and suggested that these microRNAs were involved in symptom expression. Blast analysis using the National Center for Biotechnology Information database revealed a majority of host-derived small interfering RNA sequences mapped to the previously sequenced 282 Kilobase CTV resistance gene locus of Poncirus trifoliata. Additionally, two separate P. trifoliata loci were differentially expressed; the first was centered in the intergenic and overlapping untranslated regions between CTV.11 and CTV. 12 where a Gipsy-like retrotransposone C was identified; and the second locus was between CTV.19 and CTV.20 (overlapping) genes. This project will continue for another year pending grant renewal in FY11. Focus remains examination of CTV and host genes involved in RNA silencing pathways in response to CTV infection. Research activities were monitored by the ADODR through email and telephone communication with the cooperator.