2012 Annual Report 1a.Objectives (from AD-416): Develop cost effective and highly sensitive methods to detect graft transmissible pathogens of citrus. 1b.Approach (from AD-416): Establish reliable real-time PCR detection of graft transmissible pathogens of citrus including but not limited to Pospiviroids, Hostuviroids, Spiroplasma citri, Dweet mottle virus, citrus tatter leaf virus. Establish protocols for rapid extraction of pathogen nucleic acid from target tissue. Develop genomic information of graft transmissible pathogens of citrus including pathogens which are presently uncharacterized. 3.Progress Report: The project relates to objective 4 of the parent project. Sequences of citrus pathogens are being used to develop primers for mol-PCR assays to enable easy detection of the pathogens in multiplex assays. The mol-PCR assay offers the opportunity to assay up to 200 different targets in one reaction well, and to read the results for each target using a bead-based revelation system such as the Luminex 200 instrument. Research is being conducted to develop the mol-PCR assay for detection of graft transmissible pathogens in citrus. Primers for use with the mol-PCR assay are being developed bases on sequences of citrus pathogens in GenBank.