Insertion of Agrobacterium roB gene in mango

Abstract

Transgenic mango (Mangifera indica L.) plants were regenerated from somatic embryos inoculated with a wild Agrobacterium rhizogenes strain. The bacteria were grown in Luria Bertani medium supplemented or not with acetosyringone until an optical density of 1.0-1.5 was reached. Incubation time of somatic embryos from the Kent, Haden and Madame Francis varieties in bacterial suspension was 1h, 15 and 5min. Somatic embryos were transferred into Gamborg Miller Ojima semisolid medium for 48h, and finally placed on the same medium containing cefotaxime/ carbencillin, in darkness at 27 ±1°C. In the first treatment, A. rhizogenes was eliminated after 20 washings with cefotaxime/carbencillin, only in the Kent variety; while 1-5 washes were needed for the other treatments. There were no differences between preculture treatments of A. rhizogenes with or without acetosyringone. After infection embryos formed callus, then secondary embryos and, finally, plants were regenerated but hairy roots were not induced. The presence of rolB gene in leaf tissue was confirmed by PCR. An expected band of 720bp, corresponding to rol B gen amplification, was obtained only in the transformed plants. An efficient protocol has been developed for successful production of transgenic somatic embryos and plants of the Kent variety using A. rhizogenes. Twenty transgenic clones per 0.5g of inoculated tissues with 80% embryo survival were produced. The results constitute the first report of transgenic mango plants mediated by A. rhizogenes as an alternative tool for breeding improvement of this crop.