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Transgenic callus of Nicotiana glauca stably expressing a fungal laccase maintains its growth in presence of organic contaminants

De-Jesús-García, Ramces, Folch-Mallol, Jorge Luis, Dubrovsky, Joseph G.
Plant cell, tissue, and organ culture 2019 v.138 no.2 pp. 311-324
Cauliflower mosaic virus, Nicotiana glauca, Nicotiana plumbaginifolia, Pycnoporus sanguineus, Rhizobium rhizogenes, anthraquinones, apoplast, cadmium, callus, calreticulin, catalytic activity, culture media, dyes, fungi, genes, genetic engineering, genetically modified plants, heavy metals, laccase, oxygen, phenol, phytoremediation, polluted soils, promoter regions, reaction mechanisms
Some plant species colonize mining-contaminated soils. They are adapted to harsh growth conditions and show high native phytoremediation abilities. For environmental cleanup, these species can be improved by genetic manipulation. Laccases are ligninolytic enzymes that oxidise a broad range of substrates by a radical-catalysed reaction mechanism using oxygen as the electron acceptor. Fungi of the genus Pycnoporus produce laccases with important biotechnological, industrial and environmental applications. Here we describe a successful attempt of Agrobacterium rhizogenes mediated stable transformation of Nicotiana glauca, a species naturally resistant to metal-contaminated soils. The coding region of a Pycnoporus sanguineus laccase gene was fused with the Nicotiana plumbaginifolia calreticulin apoplast targeting signal and driven by the constitutive cauliflower mosaic virus (CaMV) 35S promoter (2X35S). The obtained transgenic N. glauca callus cells secreted a recombinant fungal laccase into the growth medium and were capable to degrade an anthraquinone dye, Remazol Brilliant Blue R. Under phenol and cadmium stresses, the transgenic calli not only maintained their growth capacity but their relative growth rate was greater compared to callus transformed with empty vector. This work shows that stable expression of a fungal laccase in a plant species resistant to heavy metals presents a successful strategy that potentially can be used to combat combined organic and inorganic contamination.