Regeneration and transformation protocols for 'Comet' red raspberry were optimized with the purpose of making the Agrobacterium-mediated gene transfer system efficient for this crop. Adventitious shoot regeneration from leaf discs was improved using explants 10 mm in diameter and transferring to fresh medium at the fourth week of incubation. Additions of liquid medium to solid medium during incubation decreased regeneration and attempts to release the suppressive influence of larger shoots on initials (apical dominance) did not succeed. The presence of claforan did not affect shoot regeneration, but inoculations with Agrobacterium and the presence of kanamycin decreased regeneration moderately or considerably, respectively. The threshold for kanamycin concentration for screening for kanamycin resistant transformed raspberry tissue was 30 to 40 mg l$ sp{-1}.$ The best co-incubation interval between wild-type Agrobacterium and 'Comet' leaf discs ranged from 2 days for highly virulent strains to 3 or more days for moderate to low virulent strains. Among several wild-type strains, C58 was chosen as the most appropriate partially because a disarmed form was commercially available for use as a non-oncogenic vector for transformation of red raspberry. / The binary plasmid pBI121 containing the marker genes NPTII and GUS encoding kanamycin resistance and $ beta$-glucuronidase activity, respectively, was successfully introduced into the Agrobacterium strain LBA4404, which is a disarmed C58 derivative. Transformation of 'Comet' red raspberry was apparently achieved by inoculating leaf disc explants with LBA4404 containing pBI121. The probable integration and expression of the foreign genes into the plant cells were confirmed by screening for kanamycin resistance, GUS assays and Southern blot analyses. This transformation system appears to be effective and may be useful in further studies on red raspberry for both introduction of genes for desirable agronomic traits and basic studies of gene expression.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.69522 |
| Date | January 1993 |
| Creators | Faria, Maria José Sparça Salles de |
| Contributors | Tibellius, K. H. (advisor) |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Master of Science (Department of Microbiology.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001338490, proquestno: AAIMM87815, Theses scanned by UMI/ProQuest. |
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