Vol. 76 No: 5

Title:
Factors affecting Agrobacterium-mediated gene transfer and the selection of transgenic calli in paper shell almond (Prunus dulcis Mill.)

Authors:
PHILLIP J. AINSLEY, GRAHAM G. COLLINS and MARGARET SEDGLEY

pp: 522-528

Abstract:
Factors influencing the transfer of an intron-containing β-glucuronidase gene (GUS) to micropropagated leaf explants of the paper shell almond (Prunus dulcis Mill.) cultivars Nonpareil and Ne Plus Ultra were studied during the early steps of an Agrobacterium tumefaciens-mediated transformation procedure. The gene transfer process was evaluated by counting the number of GUS-positive leaf zones 4.d after cocultivation, as well as by counting numbers of GUS- positive calli developing on explants six weeks post-cocultivation. Of the two A. tumefaciens strains tested, EHA105 was more effective than LBA4404, and cocultivation over a 4 d period was the optimum time for both almond cultivars. The presence of acetosyringone in the bacterial, plant preculture and cocultivation media had a varied affect on gene transfer efficiency, its influence being dependent on genotype and bacterial strain. Using an optimized procedure, 1500 leaf explants of both genotypes were transformed. While no transgenic plants were recovered, PCR and Southern blot analysis confirmed the presence of the transgene and its stable integration in callus.

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