Selection system and co-cultivation medium are important determinants of Agrobacterium-mediated transformation of sugarcane
| Autor: | Melissa Kuwahata, Priya Joyce, Prakash Lakshmanan, Nicole Turner |
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| Rok vydání: | 2009 |
| Předmět: |
Rhizobiaceae
DNA Plant Paromomycin Agrobacterium Plant Science Genetically modified crops Genes Plant chemistry.chemical_compound Transformation Genetic Gene Expression Regulation Plant Genes Reporter Botany Transgenes Selectable marker biology fungi food and beverages General Medicine Agrobacterium tumefaciens Plants Genetically Modified biology.organism_classification Molecular biology Culture Media Saccharum Transformation (genetics) chemistry Cinnamates Callus Hygromycin B Genetic Engineering Agronomy and Crop Science Plasmids |
| Zdroj: | Plant Cell Reports. 29:173-183 |
| ISSN: | 1432-203X 0721-7714 |
| Popis: | A reproducible method for transformation of sugarcane using various strains of Agrobacterium tumefaciens (A. tumefaciens) (AGL0, AGL1, EHA105 and LBA4404) has been developed. The selection system and co-cultivation medium were the most important factors determining the success of transformation and transgenic plant regeneration. Plant regeneration at a frequency of 0.8-4.8% occurred only when callus was transformed with A. tumefaciens carrying a newly constructed superbinary plasmid containing neomycin phosphotransferase (nptII) and beta-glucuronidase (gusA) genes, both driven by the maize ubiquitin (ubi-1) promoter. Regeneration was successful in plants carrying the nptII gene but not the hygromycin phosphotransferase (hph) gene. NptII gene selection was imposed at a concentration of 150 mg/l paromomycin sulphate and applied either immediately or 4 days after the co-cultivation period. Co-cultivation on Murashige and Skoog (MS)-based medium for a period of 4 days produced the highest number of transgenic plants. Over 200 independent transgenic lines were created using this protocol. Regenerated plants appeared phenotypically normal and contained both gusA and nptII genes. Southern blot analysis revealed 1-3 transgene insertion events that were randomly integrated in the majority of the plants produced. |
| Databáze: | OpenAIRE |
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