Regeneration of Plants from Peach Embryo Cells Infected with a Shooty Mutant Strain of Agrobacterium

Abstract

<jats:p>Immature `Redhaven' peach [<jats:italic>Prunus persica</jats:italic> (L.) Batsch] embryos were infected with a shooty mutant strain of <jats:italic>Agrobacterium tumefaciens, tms</jats:italic>328::Tn5, which carries an octopine-type Ti plasmid with a functional cytokinin gene and a mutated auxin gene. Shoots were regenerated from embryo-derived callus that was initiated on MS medium lacking phytohormones. Shoots exhibited increased frequency of branching and were more difficult to root than the noninfected. Transcripts of the <jats:italic>tms</jats:italic>328::Tn5-cytokinin gene were detected using northern analyses of total plant RNA. Polymerase chain reaction of genomic DNA and cDNA resulted in amplification of DNA fragments specific for the cytokinin gene, as determined by restriction enzyme and Southern analyses. The concentrations of the cytokinins zeatin and zeatin riboside in the leaves of regenerated plants were on the average 51-fold higher than in leaves taken from nontransformed plants. None of the shoots or callus tissues were postive for octopine. The expression of the T-DNA encoded cytokinin gene promotes growth of peach cells in the absence of phytohormones, thus serving as a marker for transformation. In addition, this gene appears to promote morphogenesis without an auxin inductive step.</jats:p>

Evaluation of transformation in peach Prunus persica explants using green fluorescent protein (GFP) and beta-glucuronidase (GUS) reporter genes

“…The recovery of transgenic peach plants, with the rates of regeneration and transformation generally reported, is very low (Smigocki and Hammerschlag, 1991;Perez-Clemente et al, 2004). Higher rates of regeneration are necessary and, as we show in this report, Agrobacterium-mediated transformation rates must also be improved.…”

“…There are only two reports of stable transgenic peach plant production. Smigocki and Hammerschlag (1991) used a shooty mutant strain of A. tumefaciens. The several transgenic plants that were obtained expressed the bacterial cytokinin biosynthesis gene (ipt).…”

Transformation of fruit trees. Useful breeding tool or continued future prospect?

“…These armed strains interrupt an auxin synthesis gene with the transposon tn5, but the ipt gene for cytokinin synthesis remains intact. Peach plants transformed with these strains have a high production of endogenous cytokinin, greater branching and shorter internodes (Smigocki & Hammerschlag, 1991). Transformation of peach tissues with the ipt gene allowed selection of transformed shoots on a medium with low concentrations of growth regulators; these were the first transformed peaches reported.…”

Cytokinins, donor plants and time in culture affect shoot regenerative capacity of American elm leaves

“…Use of Agrobacteruim-mediated transformation has been successfully applied to woody plants (Fillatti et al 1987;McGranahan et al 1988;James et al 1989;Mante et al 1991;Smigocki & Hammerschlag 1991). Production of large numbers of shoots is critical for this transformation method, especially if transformation efficiency is low (James et al 1989).…”