Abstract
An Agrobacterium-mediated transformation protocol for Japonica rice (cv. R321), using conventional genetic vectors and explants pretreated with antinecrotic compounds is presented. We evaluated the effect of two compounds with known antioxidant activity (ascorbic acid and cysteine) and silver nitrate on the viability of stem sections taken from in vitro rice plantlets, and on their interaction with Agrobacterium tumefaciens (At 2260) containing a shuttle vector bearing the gusand bar genes. After co-culture, calli formed on the callus-induction medium were supplemented with phosphinotricin and cefotaxime; putative transgenic plants were recovered on the regeneration medium after three months. All recovered plants were challenged with the herbicide BASTA under greenhouse conditions, and some resistant individuals were analyzed using PCR and a histochemical GUS test. Southern blot analysis of several R1 transgenic plants indicated the presence of at least two intact bar gene copies per genome. Inheritance of the bar gene at the R2 generation was confirmed. Antinecrotic pretreatment of the explants provides an adequate environment for the interaction of A. tumefacienswith the plant cells, thus allowing satisfactory transformation performance without the need of super-binary vectors and hyperinfective A. tumefaciens strains.
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Enríquez-Obregón, G.A., Prieto-Samsónov, D.L., de la Riva, G.A. et al. Agrobacterium-mediated Japonica rice transformation: a procedure assisted by an antinecrotic treatment. Plant Cell, Tissue and Organ Culture 59, 159–168 (1999). https://doi.org/10.1023/A:1006307527449
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DOI: https://doi.org/10.1023/A:1006307527449