Browsing by Author "Gebauer, M"

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    Enhanced resistance to bacterial infection by Erwinia carotovora subsp atroseptica in transgenic potato plants expressing the attacin or the cecropin SB-37 genes
    (1999) Arce, P; Moreno, M; Gutierrez, M; Gebauer, M; Dell'Orto, P; Torres, H; Acuña, I; Oliger, P; Venegas, A; Jordana, X; Kalazich, J; Holuigue, L
    Blackleg and soft rot diseases, caused by the bacterium Erwinia carotovora, are among the diseases that cause important losses in culture and storage of potato. In this paper, we introduced bacterial resistance into potato, via genes encoding for proteins with antibacterial activity. For this purpose, potato clones were transformed either with the gene encoding the acidic attacin protein from Hyalophora cecropia, or with the gene encoding the cecropin analog peptide SB-37. These clones were evaluated for soft rot and blackleg resistance, after inoculation with the bacterial strain Erwinia carotovora subsp. atroseptica T7. Results reported in this paper indicate that a considerable percentage of the potato clones (15-22%) showed increased resistance to bacterial infection, revealed by reduced severity of blackleg or soft rot symptoms. Expression of the transgenes was demonstrated in some of the clones by Northern blot analysis. This is the first report indicating that. expression of the gene encoding for an attacin protein and for the cecropin SB-37 peptide in transgenic potato confers increased resistance to bacterial infection.
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    Expression of the chicken lysozyme gene in potato enhances resistance to infection by Erwinia carotovora subsp, Atroseptica
    (2000) Serrano, C; Arce-Johnson, P; Torres, H; Gebauer, M; Gutierrez, M; Moreno, M; Jordana, X; Venegas, A; Kalazich, J; Holuigue, L
    Infection of potato plants and tubers with the bacterium Erwinia carotovora subsp, atroseptica produces blackleg and soft rot diseases, which cause significant losses to crops and stored potatoes. In order to obtain resistance against this bacterium, the gene chly encoding the enzyme lysozyme from chicken was introduced into potato plants (cv. Desiree) via Agrobacterium-mediated transformation. Sixty-three and 69 transgenic potato clones were evaluated in the greenhouse for resistance to blackleg and soft rot diseases, respectively. Results reported in this paper indicate that 21%-29% of the potato clones showed increased resistance to infection by the bacterium E. c, subsp, atroseptica T7, as revealed by a reduced severity of blackleg or soft rot symptoms. Nine clones showing different levels of resistance were selected for further molecular analysis. The number of copies of the transgene integrated in the plant genome of these clones was estimated by Southern blot analysis. The level of transgene expression, detected by Northern blot analysis, correlated with the level of resistance detected in these clones.
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    Optimization of in vitro culture conditions for Pinus radiata embryos and histological characterization of regenerated shoots
    (1999) Stange, C; Prehn, D; Gebauer, M; Arce-Johnson, P
    Different in vitro culture conditions were tested on Pinus radiata organogenic embryos. Optimum shoot induction occurred at 26.1 degrees C, whereas the best elongation resulted at 21.4 degrees C. Supplements of 2.5 mg/l or 5 mg/l of BAP added to the induction media produced a similar number of regenerated shoots, which differed statistically from 1.0 mg/l of BAP and 0.025 mg/l TDZ. Addition of 10 mg/l MnSO4 to LP1/2 medium significantly increased the number and quality of in vitro regenerated shoots. The removal the apical region of shoots cultured in LP 2.5 mg/l of BAP increased the number of de novo generated shoots by 23%, compared to a control group with intact shoots. Approximately 70% of the in vitro shoots of P. radiata were of wet phenotype (hyperhydrated appearance); the rest were waxy in appearance. Histological cuts did not produce any differences in phenotypes, but scanning electronic microscopy of needles gave evidence of differences in epicuticular wax deposits. Abbreviations: LP: Quoirin and LePoivre basal medium, without plant growth regulators; LP,: LP medium + 1 mg/l BAP; LP2.5: LP medium+ 2.5 mg/l BAP; LP5 : LP medium + 5 mg/l BAP; LP1/2: LP basal medium at half strength of macroelements, 2%; commercial sugar, ammonium nitrate 100 mg/l, calcium nitrate 564.5 mg/l, hydroxyquinoleine 1.25 mg/l, MS vitamins and without plant growth regulators; LPT0.025: LP medium + 0.025 mg/l TDZ; BAP: N-6 benzylaminopurine; TDZ: Thidiazuron.
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    Stable transformation of Pinus radiata embryogenic tissue by Agrobacterium tumefaciens
    (2002) Cerda, F; Aquea, F; Gebauer, M; Medina, C; Arce-Johnson, P
    Embryogenic cultures from immature zygotic embryos of Pinus radiata seeds were established on semisolid proliferation medium with 2,4-D and BAP. Growing embryogenic masses containing embryonal cells and suspensor cells were subcultured on this media every 2 weeks. After 10 weeks, embryogenic masses (1.5 cm diameter) were transferred to a maturation medium containing ABA. Fully developed somatic embryos were obtained in this medium after 12 weeks. Embryogenic masses were genetically transformed using Agrobacterium tumefaciens. The pBI121 vector containing beta-glucuronidase (uidA) and the neomycin phosphotransferase (nptll) genes was introduced into this tissue. After co-cultivation with Agrobacterium, the embryogenic tissues were transferred to a selection media containing geneticin and carbenicillin. After 1 month of selection, histochemical assays showed extensive GUS positive activity zones in the transformed embryogenic tissues. Under light microscope, blue crystals were seen inside the embryogenic and suspensor cells, and also completely blue somatic embryos were obtained. The uidA gene was also detected by PCR analysis in genomic DNA isolated from transformed embryogenic tissues. These results indicate stable transformation of P. radiata somatic embryogenic tissues using Agrobacterium-mediated transformation.