Transgene stacking in potato using the GAANTRY system

• Published in: BMC research notes Vol. 12; no. 1; p. 457
• Main Authors: McCue, Kent F, Gardner, Ethan, Chan, Ronald, Thilmony, Roger, Thomson, James
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 25.07.2019; BioMed Central; BMC
• Subjects: Agrobacterium - genetics; Agrobacterium - metabolism; Diseases and pests; DNA, Bacterial - genetics; DNA, Bacterial - metabolism; GAANTRY; Gene Dosage; Gene Expression; Gene Transfer Techniques; Gene-stacking; Genes, Reporter; Genetic aspects; Genetically modified plants; Glucuronidase - genetics; Glucuronidase - metabolism; Growth; Luminescent Proteins - genetics; Luminescent Proteins - metabolism; Methods; Phenotype; Plant breeding; Plant Leaves - genetics; Plant Leaves - microbiology; Plants, Genetically Modified; Plasmids - chemistry; Plasmids - metabolism; Polymerase Chain Reaction - methods; Potatoes; Red Fluorescent Protein; Research Note; Site-specific recombinase; Solanum tuberosum; Solanum tuberosum - genetics; Solanum tuberosum - microbiology; Transgenes; United States; GAANTRY; Gene-stacking; Site-specific recombinase; Solanum tuberosum
• ISSN: 1756-0500; 1756-0500
• DOI: 10.1186/s13104-019-4493-8

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. 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.