ARS | Publication request: Improving regeneration and transformation for resistance to sharka in prunus

Submitted to: Acta Horticulturae
Publication Type: Proceedings
Publication Acceptance Date: December 20, 2006
Publication Date: March 10, 2007
Citation: Damiano, C., Kondakova, V., Todorovska, E., Kamenova, I., Badjiakov, I., Scorza, R., Gentile, A., Monticelli, S., Atanassov, A. 2007. Improving regeneration and transformation for resistance to sharka in prunus. Acta Horticulturae. 738:583-587.

Technical Abstract: The biotechnological approach is a promising strategy to overcome some serious diseases which affects stone fruits such as Sharka. However, members of the genus Prunus are among the most challenging with respect to in vitro organogenesis: regeneration and transformation are highly dependent on species, genotype, and source of explants. We focused our activity on the development of adventitious shoot regeneration protocols using leaves of micropropagated Prunus varieties and on the optimization of regeneration ratios, based on the hypocotyl slices strategy from mature peach seeds (Nemaguard, Missour, Montclar, Romestar, Percoco di Tursi), apricot (Sancastrese, Hatif Hatif, Vitillo, Boccuccia spinosa, Palummella), and plum (Stanley, St. Julien). The infection of hypocotyl slices of Stanley and St. Julien with Agrobacterium tumefaciens containing the plasmid pGA482GG/PPV-CP-33 and carrying the Plum pox virus (PPV) coat protein, the GUS and NPTII genes, allowed the creation of eight transgenic clones of Stanley and three transgenic clones of St. Julien. GUS activity and molecular analysis of regenerated shoots confirmed the insertion of PPV-CP and NPTII genes in the plant genome and the absence of bacterial contamination. Challenging the transgenic St. Julien clones with sap prepared from leaves of Nicotiana benthamiana infected with PPV produced no symptoms on inoculated and on post-inoculation generated leaves. DAS-ELISA results confirmed the absence of virus infection in transgenic plants.