Articles
AGROBACTERIUM TUMEFACIENS-MEDIATED TRANSFORMATION OF YELLOW PASSION FRUIT (PASSIFLORA EDULIS F. FLAVICARPA WITH THE CME-ACOL GENE
Article number
632_30
Pages
243 – 248
Language
English
Abstract
In order to improve fruit quality and shelf-life of the yellow passion fruit, experiments were carried out for the introduction of an antisense melon ACCoxidase gene (CMe-ACO1) into leaf explants via Agrobacterium tumefaciens, followed by plantlet regeneration.
For regeneration experiments, three concentrations of benzyladenine (BA), 4.44, 6.66 and 8.87 μM, were supplemented into MS medium.
Leaf explants were excised from seedlings and grafted plants and two positions of explants were compared: abaxial or adaxial face in contact with the culture medium.
Best results were obtained on a medium containing 6.66 μM BA for both kinds of explants and for both explant positions.
For seedling material, this treatment resulted in more than 60 % of explants with regenerating shoots for both positions.
Around 40 to 60 % of explants coming from grafted plants regenerated on the same medium, depending on the genotype.
Shoots were rooted on half-strength MS medium containing 2.89 μM gibberellic acid.
For transformation experiments, leaf explants were cocultured with A. tumefaciens strain EHA 101 carrying the binary vector pGA643, which includes an antisense CMe-ACO1 and a nos/nptII gene constructs.
Transformation with EHA101 carrying pBIN19 allowed the regeneration of control transgenic plants, transformed with the nptII gene only.
As indicated by PCR analysis, three plants among nine grown in the presence of kanamycin were found putatively transformed with nptII gene, and one plant among six grown under the same conditions were found putatively transformed with CMe-ACO1 gene.
For regeneration experiments, three concentrations of benzyladenine (BA), 4.44, 6.66 and 8.87 μM, were supplemented into MS medium.
Leaf explants were excised from seedlings and grafted plants and two positions of explants were compared: abaxial or adaxial face in contact with the culture medium.
Best results were obtained on a medium containing 6.66 μM BA for both kinds of explants and for both explant positions.
For seedling material, this treatment resulted in more than 60 % of explants with regenerating shoots for both positions.
Around 40 to 60 % of explants coming from grafted plants regenerated on the same medium, depending on the genotype.
Shoots were rooted on half-strength MS medium containing 2.89 μM gibberellic acid.
For transformation experiments, leaf explants were cocultured with A. tumefaciens strain EHA 101 carrying the binary vector pGA643, which includes an antisense CMe-ACO1 and a nos/nptII gene constructs.
Transformation with EHA101 carrying pBIN19 allowed the regeneration of control transgenic plants, transformed with the nptII gene only.
As indicated by PCR analysis, three plants among nine grown in the presence of kanamycin were found putatively transformed with nptII gene, and one plant among six grown under the same conditions were found putatively transformed with CMe-ACO1 gene.
Authors
M. Quoirin, L.M. Winkler, R. Ayub
Keywords
ACC oxidase, ethylene, organogenesis, PCR
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