Agrobacterium-mediated stable genetic transformation of Populus angustifolia and Populus balsamifera

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Maheshwari, Priti
Kovalchuk, Igor
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Frontiers Media
The present study demonstrates Agrobacterium tumefaciens-mediated stable genetic transformation of two species of poplar – Populus angustifolia and Populus balsamifera. The binary vector pCAMBIA-Npro-long-Luc containing the luciferase reporter gene was used to transform stem internode and axillary bud explants. Putative transformants were regenerated on selection-free medium using our previously established in vitro regeneration method. Explant type, genotype, effect of pre-culture, Agrobacterium concentration, a time period of infection and varying periods of co-culture with bacteria were tested for the transformation frequency. The highest frequency of transformation was obtained with stem internode explants pre-cultured for 2 days, infected with Agrobacterium culture at the concentration of OD600 D 0.5 for 10 min and co-cultivated with Agrobacterium for 48 h. Out of the two genotypes tested, P. balsamifera exhibited a higher transformation rate in comparison to P. angustifolia. The primary transformants that exhibited luciferase activity in a bioluminescence assay under the CCD camera when subjected to polymerase chain reaction and Southern blot analysis revealed a stable single-copy integration of luc in their genomes. The reported protocol is highly reproducible and can be applied to other species of poplar; it will also be useful for future genetic engineering of one of the most important families of woody plants for sustainable development.
Sherpa Romeo green journal: open access
Agrobacterium tumefaciens , Luciferase , Poplar , Populus angustifolia , Populus balsamifera , Regeneration , Stable transformation
Maheshwari, P., & Kovalchuk, I. (2016). Agrobacterium-mediated stable genetic transformation of Populus angustifolia and Populus balsamifera. Frontiers in Plant Science, 7(296). doi:10.3389/fpls.2016.00296