The efficient production of marker-free transgenic plants is still a challenge in most fruit species even though such plants are a necessary component of many "new breeding technologies", particularly cis- and intragenesis. Marker-free plant production is also necessary for the successive stacking of genes in an elite fruit transgenic line. Here, we used a R/Rs site-specific recombinase that is post-translationally regulated by dexamethasone through fusion with a ligand-binding domain for this hormone, and a bi-functional selectable marker gene coding for a cytosine deaminase/neomycin transferase (codA-nptII) protein; this enabled a first step of positive kanamycin selection, followed by a second step of negative 5-fluorocytosine selection. The aim of our study was to optimize this system on the apple cv. Galaxy and on the pear cv. Conference by conducting a detailed study of the effects of dexamethasone and 5-fluorocytosine treatments, and by comparing an early versus a delayed selection strategy. We were able to produce marker-free transgenic pear plants for the first time, and confirm the feasibility of producing marker-free transgenic apple plants using a chemically inducible recombinase system. We recommend the use of an early selection strategy for the pear cv. Conference and a delayed selection strategy for the apple cv. Galaxy
The biological effects of exposure to electromagnetic fields due to wireless technologies and connected devices are a subject of particular research interest. Ultrashort high‐amplitude electromagnetic field pulses delivered to biological samples using immersed electrodes in a dedicated cuvette have widely demonstrated their effectiveness in triggering several cell responses including increased cytosolic calcium concentration and reactive oxygen species (ROS) production. In contrast, the effects of these pulses are poorly documented when electromagnetic pulses are delivered through an antenna. Here we exposed Arabidopsis thaliana plants to 30,000 pulses (237 kV m−1, 280 ps rise‐time, duration of 500 ps) emitted through a Koshelev antenna and monitored the consequences of electromagnetic fields exposure on the expression levels of several key genes involved in calcium metabolism, signal transduction, ROS, and energy status. We found that this treatment was mostly unable to trigger significant changes in the messenger RNA accumulation of calmodulin, Zinc‐Finger protein ZAT12, NADPH oxidase/respiratory burst oxidase homolog (RBOH) isoforms D and F, Catalase (CAT2), glutamate‐cystein ligase (GSH1), glutathione synthetase (GSH2), Sucrose non‐fermenting‐related Kinase 1 (SnRK1) and Target of rapamycin (TOR). In contrast, Ascorbate peroxidases APX‐1 and APX‐6 were significantly induced 3 h after the exposure. These results suggest that this treatment, although quite strong in amplitude, is mostly ineffective in inducing biological effects at the transcriptional level when delivered by an antenna. © 2023 The Authors. Bioelectromagnetics published by Wiley Periodicals LLC on behalf of Bioelectromagnetics Society.
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