McCue Kent F, Gardner Ethan, Chan Ronald, Thilmony Roger, Thomson James
Crop Improvement and Genetics, Western Regional Research Center, USDA-ARS, Albany, CA, USA.
BMC Res Notes. 2019 Jul 25;12(1):457. 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.
GAANTRY（利用重组酶技术通过核酸转移在农杆菌中进行基因组装）是一种灵活且有效的系统，用于在农杆菌毒性质粒转移DNA（T-DNA）中稳定堆叠多个基因。我们检测了GAANTRY发根农杆菌ArPORT1“10堆叠”菌株产生转基因马铃薯植株的能力。
将28.5千碱基的10堆叠T-DNA导入Lenape马铃薯植株，转化效率为32%。分子和表型特征证实，7个测试的独立转基因株系中有6个携带了完整的所需构建体，表明GAANTRY 10堆叠菌株可用于基于组织培养的愈伤组织转化方法，以高效产生转基因马铃薯植株。使用液滴数字PCR分析表明，大多数已鉴定的事件携带10堆叠转基因的一个或两个拷贝，并且转基因植株中不存在来自T-DNA外部的“骨架”DNA。这些结果表明，GAANTRY系统能高效产生具有大量堆叠转基因构建体的高质量转基因马铃薯植株。
