Challenging Transgenic ‘W. Murcott’ (Nadorcott) Mandarin Containing Xa21 Gene Against Citrus Canker Pathogen

Genetic transformation offers an attractive alternative to the conventional genetic improvement of Citrus sp. Genetic transformation of several Citrus genotypes has been achieved by co-cultivating different explants (mainly juvenile) with Agrobacterium tumefaciens. However, mandarin is considered to be the most difficult Citrus sp. for transformation using the Agarobacterium-mediated system. ‘W. Murcott’ mandarin (a hybrid of ‘Murcott’ and an unknown pollen-parent) is a commercially important cultivar grown in many regions around the world. Protoplast-GFP transformation opens a new avenue to produce transgenic plants especially from seedless or other polyembryonic cultivars not amenable for the Agraobacterium-mediated system. We have produced 10 transgenic lines of ‘W. Murcott’ with plasmid DNA (pAO3), encoding the non-destructive selectable marker GFP (Green Fluorescent Protein) gene and the cDNA of the Xa21 Xanthomonas resistance gene from rice using protoplast/GFP transformation system. All plants are showing stable GFP expression all the time. PCR analysis revealed the presence of the Xa21 and the GFP genes in the transgenic plants. Molecular characterization including Southern blot analysis and Western blot analysis are showing integration and expression of the transgene in the generated transgenic plants. Q-PCR are showing different level of Xa21 gene expression among the generated transgenic lines. Transgenic plants will be evaluated against citrus canker pathogen by detached leaf assay (DLA), and by syringe inoculation of attached leaves. Results from these assays will be presented at the meeting.