Zinc tolerance and accumulation in stable cell suspension cultures and in vitro regenerated plants of the emerging model plant Arabidopsis halleri (Brassicaceae)
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Arabidopsis halleri is increasingly employed as a model plant for studying heavy metal hyperaccumulation. With the aim of providing valuable tools for studies on cellular physiology and molecular biology of metal tolerance and transport, this study reports the development of successful and highly efficient methods for the in vitro regeneration of A. halleri plants and production of stable cell suspension lines. Plants were regenerated from leaf explants of A. halleri via a three-step procedure: callus induction, somatic embryogenesis and shoot development. Efficiency of callus proliferation and regeneration depended on the initial callus induction media and was optimal in the presence of 1 mg L−1 2,4-dichlorophenoxyacetic acid, and 0.05 mg L−1 benzylaminopurine. Subsequent shoot and root regeneration from callus initiated under these conditions reached levels of 100% efficiency. High friability of the callus supported the development of cell suspension cultures with minimal cellular aggregates. Characterization of regenerated plants and cell cultures determined that they maintained not only the zinc tolerance and requirement of the whole plant but also the ability to accumulate zinc; with plants accumulating up to 50.0 μmoles zinc g−1 FW, and cell suspension cultures 30.9 μmoles zinc g−1 DW. Together this work will provide the experimental basis for furthering our knowledge of A. halleri as a model heavy metal hyperaccumulating plant.
KeywordsArabidopsis halleri Heavy metal hyperaccumulation Cell suspensions Zinc tolerance Plant regeneration
MS medium supplemented with 0.5 mg L−1 2,4- dichlorophenoxyacetic acid (2,4-D) and 0.5 mg L−1 kinetin
MS medium supplemented with 1 mg L−1 2,4-D, and 0.05 mg L−1 benzylaminopurine, BA
MS medium supplemented with 0.5 mg L−1 2,4-D, 0.1 mg L−1 BA
0.5× MS medium supplemented with 0.3 mg L−1 2,4-D
Murashige and Skoog medium supplemented with 10 mg L−1 thiamine hydrochloride, 1 mg L−1 nicotinic acid, 1 mg L−1 pyridoxine hydrochloride, 100 mg L−1 myo-inositol, 3% sucrose and 1.5% bacteriological agar (pH 5.7 ± 0.1)
We thank Dr. Ute Krämer from the Heidelberg Institute of Plant Sciences, University of Heidelberg for kindly providing A. halleri seeds. This work was supported by funding from CONACYT, grants 57685 to RV-E and 49765 to BJB; and DGAPA IN221308 to RV-E.
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