As a first step to the establishment of a genetic transformation protocol for olive somatic embryos obtained from the seeds of cv. 'Picual', the efficiencies of different aminoglycoside antibiotics as selective agents to be used with the nptII marker gene, and the particle bombardment technique for transient transformation have been evaluated. Among the three antibiotics tested, paromomycin and kanamycin showed a similar inhibitory effect and, at 200 mg l -1 , both of them impaired callus growth after 8 weeks of culture. However, when isolated embryos were cultured in the presence of these antibiotics, a 20% of the embryos still remained viable at 400 mg l -1 . Neomycin was discarded as a selective agent since it showed only a moderate toxic effect. Contrary to solid medium, when olive callus was cultured in liquid medium supplemented with different paromomycin concentrations for 3 weeks, the callus growth was impaired at the lowest antibiotic concentration, 3 mg l -1 . Best conditions for transient transformation of olive callus using PDS-1000/He system were a 6 cm target distance and a 900 psi bombardment pressure. pCGUD1 plasmid, containing the gus gene under the control of sunflower ubiquitin promoter yielded a significantly higher number of gus expression areas per bombarded explant than pGUSINT or pJGUS5 plasmids, where the gus gene is driven by CaMV35S promoter or CaMV35S with enhancer, respectively. Almost 45% of bombarded explants showed gus expression 12 weeks after bombardment.

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Avocado embryogenic cell cultures can be classified into two groups based on their morphology when cultured on a medium containing auxin: somatic embryo (SE) and proembryonic masses (PEM) type cultures. The calli of SE-type cell lines are able to go through the maturation process, whereas the calli of PEM cell lines rarely mature. We have investigated four independent avocado cell cultures (two SE and two PEM). The aim of this study was to link the differential regeneration capacity of the four cell cultures to a proteomic pattern and to gain insight into the regeneration capacity. A 2D-DIGE analysis followed by a blind multivariate analysis was able to separate the two SE lines from the PEM lines indicating that the protein profiles of SE and PEM calli are different. Based on the variable importance, that is, the differential protein pattern, we hypothesize that the regeneration capacity in avocado is correlated to the ability to overcome the physicochemical stress stimuli associated with the in vitro culture. Our identical culture conditions do not seem to trigger an appropriate response in PEM lines.

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