Summary: The breeding of plantation forestry trees for the possible afforestation of marginal land would be one approach to addressing global warming issues. Here, we developed novel transgenicEucalyptustrees (Eucalyptus camaldulensisDehnh.) harbouring anRNA‐Binding‐Protein(McRBP) gene derived from a halophyte plant, common ice plant (Mesembryanthemum crystallinumL.). We conducted screened‐house trials of the transgenicEucalyptususing two different stringency salinity stress conditions to evaluate the plants’ acute and chronic salt stress tolerances. Treatment with 400 mM NaCl, as the high‐stringency salinity stress, resulted in soil electrical conductivity (EC) levels >20 mS/cm within 4 weeks. With the 400 mM NaCl treatment, >70% of the transgenic plants were intact, whereas >40% of the non‐transgenic plants were withered. Treatment with 70 mM NaCl, as the moderate‐stringency salinity stress, resulted in soilEClevels of approx. 9 mS/cm after 2 months, and these salinity levels were maintained for the next 4 months. All plants regardless of transgenic or non‐transgenic status survived the 70 mM NaCl treatment, but after 6‐month treatment the transgenic plants showed significantly higher growth and quantum yield of photosynthesis levels compared to the non‐transgenic plants. In addition, the salt accumulation in the leaves of the transgenic plants was 30% lower than that of non‐transgenic plants after 15‐week moderate salt stress treatment. There results suggest thatMcRBPexpression in the transgenicEucalyptusenhances their salt tolerance both acutely and chronically.

Novel transgenic Eucalyptus camaldulensis trees expressing the bacterial choline oxidase A (codA) gene by the Cauliflower mosaic virus (CaMV) 35S promoter and the Arabidopsis thaliana heat shock protein (HSP) terminator was developed. To evaluate the codA transcription level and the metabolic products and abiotic stress tolerance of the transgenic trees, a six-month semi-confined screen house cultivation trial was conducted under a moderate-stringency salt-stress condition. The transcription level of the CaMV 35S promoter driven-codA was more than fourfold higher, and the content of glycine betaine, the metabolic product of codA, was twofold higher, with the HSP terminator than with the nopaline synthase (NOS) terminator. Moreover, the screen house cultivation revealed that the growth of transgenic trees under the salt stress condition was alleviated in correlation with the glycine betaine concentration. These results suggest that the enhancement of codA transcription by the HSP terminator increased the abiotic stress tolerance of Eucalyptus plantation trees.

Under the Japanese biosafety regulatory framework for transgenic plants, data for assessing a transgenic plant’s impact on biodiversity must be submitted in order to obtain approval for a confined field trial. We recently reported the development of four novel transgenic Eucalyptus camaldulensis clones expressing the bacterial choline oxidase A (codA) gene, i.e., codAH-1, codAH-2, codAN-1, and codAN-2, and evaluated their abiotic tolerance by semiconfined screen house trial cultivation. Here we evaluated the impacts of the transgenic E. camaldulensis clones on productivities of harmful substances from those clones to affect soil microorganisms and/or other plants in the environment. A comparison of the assessment data between the transgenic trees and non-transgenic comparators showed no significant difference in potential impacts on biodiversity. The results contribute to sound-science evidence ensuring substantial equivalence between transgenic and non-transgenic E. camaldulensis.

We have known that plant defensins have the main roles in biotic stress, especially antifungal activities. In addition, their functions on abiotic stress tolerance have been demonstrated on previous studies, but that information was still limited. Ca-AFP, a plant defensin gene derived from chickpea, was transformed into tobacco leaf disks by Agrobacterium method. After 16 weeks on selection media containing 15 mg/L hygromycin, some regeneration shoot lines were obtained. Four of them showed the positive bands on hpt gene and Ca-AFP gene by PCR screening. These lines were applied on in vitro evaluation of dehydration tolerance using rotary liquid culture, and the water-deficit stress was caused by 2% (w/v) PEG 6000. These four transgenic lines had better growth of adventitious roots (number of adventitious roots, total adventitious root length, and average of root length) and higher rate of biomass increase than the non-transgenic line. The result illustrated that the expression of Ca-AFP gene could mitigate the negative effect of water-deficit stress on transgenic tobacco lines under the in vitro stress condition. These transgenic tobacco lines are potential for water-deficit stress trials on greenhouse in the future.