1. Transgene stacking in potato using the GAANTRY system.
- Author
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McCue KF, Gardner E, Chan R, Thilmony R, and Thomson J
- Subjects
- Agrobacterium metabolism, DNA, Bacterial metabolism, Gene Dosage, Gene Expression, Genes, Reporter, Glucuronidase genetics, Glucuronidase metabolism, Luminescent Proteins genetics, Luminescent Proteins metabolism, Phenotype, Plant Leaves genetics, Plant Leaves microbiology, Plants, Genetically Modified, Plasmids chemistry, Polymerase Chain Reaction methods, Solanum tuberosum microbiology, Red Fluorescent Protein, Agrobacterium genetics, DNA, Bacterial genetics, Gene Transfer Techniques, Plasmids metabolism, Solanum tuberosum genetics, Transgenes
- Abstract
Objective: GAANTRY (Gene Assembly in Agrobacterium by Nucleic acid Transfer using Recombinase technologY) is a flexible and effective system for stably stacking multiple genes within an Agrobacterium virulence plasmid Transfer-DNA (T-DNA). We examined the ability of the GAANTRY Agrobacterium rhizogenes ArPORT1 '10-stack' strain to generate transgenic potato plants., Results: The 28.5 kilobase 10-stack T-DNA, was introduced into Lenape potato plants with a 32% transformation efficiency. Molecular and phenotypic characterization confirmed that six of the seven tested independent transgenic lines carried the entire desired construct, demonstrating that the GAANTRY 10-stack strain can be used can be used in a tissue culture-based callus transformation method to efficiently generate transgenic potato plants. Analysis using droplet digital PCR showed that most of the characterized events carry one or two copies of the 10-stack transgenes and that 'backbone' DNA from outside of the T-DNA was absent in the transgenic plants. These results demonstrate that the GAANTRY system efficiently generates high quality transgenic potato plants with a large construct of stacked transgenes.
- Published
- 2019
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