Background: To create an ideotype woody bioenergy crop with desirable growth and biomass properties, we utilized the viral 2A-meidated bicistronic expression strategy to express both <
em>
PtrMYB3<
/em>
(<
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MYB46<
/em>
ortholog of <
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Populus trichocarpa<
/em>
, a master regulator of secondary wall biosynthesis) and <
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PdGA20ox1<
/em>
(a GA20-oxidase from <
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Pinus densiflora<
/em>
that produces gibberellins) in wood-forming tissue (i.e., developing xylem). Results: Transgenic <
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Arabidopsis<
/em>
plants expressing the gene construct DX15::PdGA20ox1-2A-PtrMYB3 showed a significant increase in both stem fresh weight (threefold) and secondary wall thickening (1.27-fold) relative to wild-type (WT) plants. Transgenic poplars harboring the same gene construct grown in a greenhouse for 60�days had a stem fresh weight up to 2.6-fold greater than that of WT plants. In a living modified organism (LMO) field test conducted for 3�months of active growing season, the stem height and diameter growth of the transgenic poplars were 1.7- and 1.6-fold higher than those of WT plants, respectively, with minimal adverse growth defects. Although no significant changes in secondary wall thickening of the stem tissue of the transgenic poplars were observed, cellulose content was increased up to 14.4 wt% compared to WT, resulting in improved saccharification efficiency of the transgenic poplars. Moreover, enhanced woody biomass production by the transgenic poplars was further validated by re-planting in the same LMO field for additional two growing seasons. Conclusions: Taken together, these results show considerably enhanced wood formation of our transgenic poplars, with improved wood quality for biofuel production.