1a.Objectives (from AD-416):
1. Collaborate in developing a sensitive assay for Cacao swollen-shoot virus for diagnosis of virus infection in leaves suitable for scaling up to testing large sample numbers.
2. Collaborate in generating haploids via anther culture for cacao as well as cell cultures for genetic studies. Using double haploids will increase our knowledge of the genetics of these species since selected parents can be used to design new breeding populations. Plant cell cultures can be used too as sources of mitochondrial DNA for genomic studies.
1b.Approach (from AD-416):
The assays for cacao swollen-shoot virus will be developed using a real time reverse-transcriptase polymerase chain reaction approach, with primers specific to each virus. Primer design will aim at detecting the various genome variants known to occur in cacao and avocado. These assays can be later used at the ARS, and in regions of interest to the U.S., to assess specific tree populations, compare virus titers among trees and among different organs within a tree.
A plant tissue culture approach will be used to develop haploids and double haploids in cacao. Various nutritional culture media, plant growth regulators and environmental growth conditions will be tested during the development of optimal protocols for the plant cultures.
The collaboration between St. Thomas University and ARS in Miami should result in the development of molecular methods for the quick and sensitive detection of cacao swollen-shoot virus. These assays can be later adapted to of high throughput methods and used at the ARS, and in regions of interest to the U.S., to test specific tree populations, compare virus titers among trees and among different organs within a tree. The identification of the best tissue for fast and reliable virus diagnosis will allow early detection and removal of infected trees to prevent spread of infection.
Plant tissue culture protocols for production of haploid cultures and as sources of cacao mitochondrial DNA will be developed.
This research is directly related to the inhouse objective 2. Conduct expanded national and international field trials and phenotypic evaluations of cacao germplasm and breeding populations.
Developed a senstive assay for Cacao Swollen Shoot Virus (CSSV) that can be performed in cocoa producing countries.