ISHS Acta Horticulturae 455: V International Mango Symposium
TRANSFORMATION OF MANGO SOMATIC EMBRYOS |
| Authors: | A. Cruz-Hernández, Miguel A. Gómez-Lim, R. E. Litz |
| Keywords: | Mangífera indica, somatic embryogenesis, genetic transformation, fruit ripening, antisense RNA |
| Abstract:
Genetic manipulation of mango with appropiate genes could help understand several developmental processes and produce fruits with desirable traits for marketing In this work we report the construction of binary vectors with antisense genes of ripening-related genes and their transfer to somatic proembryos of mango by coculture with Agrobacterium tumefasciens. The cDNAs of ACC oxidase, ACC synthase and alternative oxidase were isolated from a mesocarp library by in situ hybridization and PCR reactions. 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 promotor. The constructs were then transferred to the LBA4404 Agrobacterium tumefaciens strain by electroporation. Somatic proembryos of mango ('Hindi') were gently macerated and co-cultivated for three days with a acetosyringone-induced culture of Agrobacterium carrying the vectors. The cultures were selected on solid medium supplemented with 100 mg/l kanamycin for four months. Transformed somatic embryos capable of growing on selection medium were assayed for GUS reaction and tested by npt II-PCR reaction and Southern blot hybridization. Proembryos were transferred to liquid medium with 200 mg/l kanamycin and finally transferred to maturation medium containing 200 mg/l kanamycin for transformed somatic embryo recovery. | |
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