Abstract
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 = 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.
Highlights
Poplars are deciduous trees of the willow family (Salicaceae) with over 30 species distributed in the temperate areas of the Northern hemisphere that differ in leaf size and shape as well as in the color of the bark, etc
Luciferase reporter gene expression was employed to monitor the transient expression for studying the effect of various factors for the optimization of transformation of P. angustifolia and
No significant difference in the transient expression of the luciferase reporter was recorded between stem internodes and axillary bud explants pre-cultured for a period of 0– 7 days in both P. angustifolia and P. balsamifera
Summary
Poplars are deciduous trees of the willow family (Salicaceae) with over 30 species distributed in the temperate areas of the Northern hemisphere that differ in leaf size and shape as well as in the color of the bark, etc. They have a short life span but are one of the fastest growing temperate trees – they can grow 5–10 feet per year (Stanturf et al, 2001). They are most important tree species in the snowboards and musical instrument industry Their wood is used for panel painting which includes the famous Mona Lisa. Poplars are very desirable as of lately for biofuel production because
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