SummaryPlant grafting is an important technique for horticultural and silvicultural production. However, many rootstock plants suffer from undesirable lateral bud outgrowth, low grafting success rates or poor rooting. Here, we used a root‐predominant gene promoter (SbUGT) to drive the expression of a tryptophan‐2‐monooxygenase gene (iaaM) from Agrobacterium tumefaciens to increase auxin levels in tobacco. The transgenic plants, when used as a rootstock, displayed inhibited lateral bud outgrowth, enhanced grafting success rate and improved root initiation. However, root elongation and biomass of SbUGT::iaaM transgenic plants were reduced compared to those of wild‐type plants. In contrast, when we used this same promoter to drive CKX (a cytokinin degradation gene) expression, the transgenic tobacco plants displayed enhanced root elongation and biomass. We then made crosses between the SbUGT::CKX and SbUGT::iaaM transgenic plants. We observed that overexpression of the CKX gene neutralized the negative effects of auxin overproduction on root elongation. Also, the simultaneous expression of both the iaaM and CKX genes in rootstock did not disrupt normal growth and developmental patterns in wild‐type scions. Our results demonstrate that expression of both the iaaM and CKX genes predominantly in roots of rootstock inhibits lateral bud release from rootstock, improves grafting success rates and enhances root initiation and biomass.

SummaryFlowerless trait is highly desirable for poplar because it can prevent pollen‐ and seed‐mediated transgene flow. We have isolated the second intron of PTAG2, an AGAMOUS (AG) orthologue from Populus trichocarpa. By fusing this intron sequence to a minimal 35S promoter sequence, we created two artificial promoters, fPTAG2I (forward orientation of the PTAG2 intron sequence) and rPTAG2I (reverse orientation of the PTAG2 intron sequence). In tobacco, expression of the β‐glucuronidase gene (uidA) demonstrates that the fPTAG2I promoter is non‐floral‐specific, while the rPTAG2I promoter is active in floral buds but with no detectable vegetative activity. Under glasshouse conditions, transgenic tobacco plants expressing the Diphtheria toxin A (DT‐A) gene driven by the rPTAG2I promoter produced three floral ablation phenotypes: flowerless, neuter (stamenless and carpel‐less) and carpel‐less. Further, the vegetative growth of these transgenic lines was similar to that of the wild‐type plants. In field trials during 2014 and 2015, the flowerless transgenic tobacco stably maintained its flowerless phenotype, and also produced more shoot and root biomass when compared to wild‐type plants. In poplar, the rPTAG2I::GUS gene exhibited no detectable activity in vegetative organs. Under field conditions over two growing seasons (2014 to the end of 2015), vegetative growth of the rPTAG2I::DT‐A transgenic poplar plants was similar to that of the wild‐type plants. Our results demonstrate that the rPTAG2I artificial promoter has no detectable activities in vegetative tissues and organs, and the rPTAG2I::DT‐A gene may be useful for producing flowerless poplar that retains normal vegetative growth.

Root sprouts-the formation of new shoots from roots-is an important mechanism for local gene flow from poplar (Populus spp). An effective strategy to reduce root sprout formation could therefore help to ensure containment during field research and commercial deployment of poplar when grown as exotic or transgenic forms. We used a flavonoid glycosyltransferase gene promoter from Scutellaria barbata (SbUGT) to drive the expression of AtCKX2, a cytokinin oxidase from Arabidopsis that converts active to inactive cytokinins in roots of poplar. In the greenhouse, SbUGT::AtCKX2 transgenic plants exhibited a similar shoot growth habit, but had enhanced root growth and fewer root sprouts, compared to the wild-type control and transgenic events with low transgene expression in roots. Under field conditions, the transgenic trees also had similar growth habits and stem growth rates that were not statistically different from wild-type trees over 3 years. Removal of trunks generally induced high rates of root sprouting; however, in selected SbUGT::AtCKX2 transgenic poplar events there was an absence or fewer root sprouts compared to wild-type trees, consistent with the greenhouse results. Our study demonstrates that the SbUGT::AtCKX2 gene can effectively inhibit root sprouting of poplar trees under field conditions, and thus may provide a useful tool to address concerns associated with root-sprouting-mediated transgene spread.