Transformation of mango somatic embryos.
Cruz-Hernández A., Gómez-Lim M., Litz R. E.
Author Affiliation: CINVESTAV-Unidad Irapuato, Apartado Postal 629, Irapuato, Gto., Mexico.
: 292-298
Abstract : Binary vectors were constructed with antisense genes of ripening-related genes and were transferred to somatic proembryos of mangoes (Mangifera indica) by coculture with Agrobacterium tumefaciens. The cDNAs of ACC oxidase, ACC synthase and alternative oxidase were isolated from a mesocarp library by in situ hybridization and the polymerase chain reaction (PCR). These sequences were at least 65% homologous with the reported sequences in other plant species. The cDNAs were cloned in the antisense orientation in the pBI121 binary vector under the control of the 35S constitutive promoter. The constructs were then transferred to A. tumefaciens LBA4404 by electroporation. Somatic proembryos of mango cv. Hindi were gently macerated and co-cultivated for 3 days with an acetosyringone-induced culture of Agrobacterium carrying the vectors. The cultures were selected on solid medium supplemented with 100 mg kanamycin/litre for 4 months. Transformed somatic embryos capable of growing on selection medium were assayed for GUS reaction, and were tested by nptII-PCR and Southern blot hybridization. Proembryos were transferred to liquid medium with 200 mg kanamycin/litre and finally transferred to maturation medium containing 200 mg kanamycin/litre for transformed somatic embryo recovery.