Journal
CURRENT GENETICS
Volume 48, Issue 6, Pages 380-388Publisher
SPRINGER
DOI: 10.1007/s00294-005-0034-1
Keywords
Glomerella graminicola; green fluorescent protein; GFP; binary vector
Categories
Ask authors/readers for more resources
We have developed an Agrobacterium tumefaciens-mediated transformation (ATMT) protocol for the plant pathogenic fungus Colletotrichum graminicola, the cause of anthracnose leaf blight and stalk rot of corn. The ATMT results in higher transformation efficiencies than previously available polyethylene glycol-mediated protocols, and falcate spores can be used instead of protoplasts for transformation. Various experimental parameters were tested for their effects on transformation efficiencies. The parameters with the greatest influence were the A. tumefaciens strain used and the Ti-plasmid it carried, the ratio of bacterium to fungus during cocultivation, and the length of cocultivation. Southern analysis demonstrated that most transformants (80%) contained tandem integrations of plasmid sequences, and at least 36% had integrations at multiple sites in the genome. In a majority of cases (70%), the whole Ti-plasmid, and not just the T-DNA, had integrated as a series of tandem repeats. Tandem integrations, especially of the whole plasmid, make it difficult to rescue DNA from both flanks of the integrations with standard PCR-based approaches. Thus, ATMT may be unsuitable for insertional mutagenesis of C. graminicola without further modification.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available