Genetic Transformation and Gene Editing: Currently the MCF is able to produce mature citrus transgenics using a standard Agrobacterium-mediated transformation protocol and a biolistic protocol. Biolistic transformation with minimal cassettes is intended for the production of single copy transgenics without plant pest sequences, which avoids one step in the regulatory hurdle. Independent transgenic events will be produced and transgenics reconfirmed by GUS or GFP assays and/or PCR for the gene of interest. The mature scion genotypes currently amenable to transformation include Valencia, Hamlin, Pineapple, and Valencia-like B770. B770 is a new Valencia-like scion cultivar that is harvested in December & January. Recently two additional higher yielding sweet orange varieties from the UF Plant Improvement Team (OLL8 and Vernia) and a new red grapefruit were introduced and are being tested in tissue culture and transformation. We have also introduced a Mandarin and a Pomelo cultivar for tissue culture and transformation tests. Mature rootstock cvs.Carrizo, Swingle, and nematode resistant Kuharski can also be used to produce transgenics if seed is desired. We are also working on a DNA free gene editing protocol.
Procedures for Participation: To submit genetic constructs to the MCF, the following is required:
1. Evidence that the transgene(s) confers resistance/tolerance to a pest/pathogen (our priority is HLB), or confers another value-added trait of importance.
2. Source of the transgene and whether it is licensed to other parties.
3. The sequence of the transgene.
4. The identity of the binary or biolistic vector, a plasmid map delimiting the location and size of genetic elements, plasmid size, and location of restriction sites.
5. It is preferable to receive vector DNA rather than vectors mobilized in Agrobacterium. Gene fragments are also acceptable and the MCF will perform the sub-cloning (see Options for Transgenic Plants).
6. Additional information may be requested, depending on the submitted transgene or other special requests.
7. If you are a UF PI, please provide your UF EHS/IBC rDNA registration no.