Transcriptional enhancement of a bacterial <i>choline oxidase A</i> gene by an <i>HSP</i> terminator improves the glycine betaine production and salinity stress tolerance of <i>Eucalyptus camaldulensis</i> trees

Abstract

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.

Development and evaluation of novel salt‐tolerantEucalyptustrees by molecular breeding using anRNA‐Binding‐Proteingene derived from common ice plant (Mesembryanthemum crystallinumL.)

“…The genetic backgrounds of the transgenic E. camaldulensis generated by the method used in this study were heterozygotic, because hypocotyls of the seedlings derived from the bulk seeds were used as the explants in the transformation experiments. We also observed similar morphological characteristics in the transgenic E. camaldulensis harbouring another transgene but generated around the same time (Tran et al ., ). From these facts, we considered that the morphological characteristics of 2‐5‐4 and 2‐5‐7 were not caused by McRBP expression but rather by polymorphism within the bulk seeds.…”

“…It was reported that a transgenic E. camaldulensis harbouring a codA gene—which was derived from the soil bacterium Arthrobacter globiformis (Ikuta et al ., ) and encodes an enzyme related to the synthesis of glycine betaine, an osmoprotectant (Ashraf and Foolad, ; Chen and Murata, ; Giri, ; Kurepin et al ., )—also survived and showed a significant reduction of growth decline due to salt stress compared with the non‐transgenic E. camaldulensis lines (Tran et al ., ). Relative growth under 6‐month salt treatment was higher in the McRBP transgenic lines than the codA transgenic lines—i.e., the relative growths of the McRBP , codA , and non‐transgenic lines were in the ranges of 8.0–10.0, 6.8–7.7, and 4.6–6.0, respectively (Figure c; Tran et al ., ). These results would suggest that McRBP is superior to codA as a transgene conferring chronic moderate stress‐tolerance to E. camaldulensis , but other factors, such as the transcriptional levels and the insertion positions of their transgenes, should also be considered.…”

“…From these situation, three independent non‐transgenic lines of E. camaldulensis (cam2, cam6, and CML2) were used as the controls (Yu et al ., ). Propagation of plantlets used for the screen house trials were performed by the stem cutting method described in our previous report (Tran et al ., ). The plantlets were pre‐cultivated in screen house for approximated 9 months and used for the trials from May 2017.…”

“…The plantlets were pre‐cultivated in screen house for approximated 9 months and used for the trials from May 2017. The detail condition on the screen house cultivation was described in the previous reports (Tran et al ., ).…”

“…We have speculated that the breeding of abiotic stress‐tolerant plantation trees could be one way to combat global climate change and contribute to the sustainable development of humankind. We have conducted molecular breeding of abiotic stress‐tolerant Eucalyptus trees using genetic resources of abiotic stress‐tolerant genes from halobacteria and/or halophytes (Kikuchi et al ., ; Matsunaga et al ., ; Oguchi et al ., ; Tran et al ., ; Yu et al ., , ,b,c,d). Mangrin was found to be a candidate salt‐tolerant gene isolated from a cDNA library from suspension‐cultured cells of a mangrove plant, Bruguiera sexangula by a functional screening for cDNA encoding proteins essential for salt tolerance (Yamada et al ., ).…”