Tetraploid plants of Zizyphus jujuba Mill. cv. Zhanhua were obtained with in vitro colchicine treatment. Shoot tips from in vitro-grown plants were treated with five different concentrations of colchicine (0.01, 0.03, 0.05, 0.1, 0.3%) in liquid MS medium (Murashige and Skoog 1962), and shaken (100 rpm) at 25 degrees C in darkness for 24, 48, 72 or 96 h, respectively. Tetraploids were obtained at a frequency of over 3% by using 0.05% colchicine (48 h, 72 h) and 0.1% colchicine (24 h, 48 h) treatment as determined by flow cytometry. Cytological and morphological evidence confirmed the results of flow cytometric analysis. The chromosome number of diploid plants was 24 and that of tetraploid plants was 48. The stomata sizes of tetraploid plants were significantly larger than those of diploid plants, while the frequency of stomata were reduced significantly. Similarly, the chloroplast number of guard cells of tetraploid plants increased significantly. The selected tetraploid plants were grafted onto mature trees of Z. jujuba Mill. cv. Zhanhua in the field, resulted in thicker stems, rounder and succulent leaves, larger flowers and a delay in florescence time (3-4 days later) than diploid plants.

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Conditions for Agrobacterium-mediated transformation of wheat (Triticum aestivum L.) were defined using wheat suspension cells as a model system and green fluorescent protein (GFP) as a visual marker. Different strains of Agrobacterium tumefaciens were compared using established wheat cell suspension cultures, where the frequency of cell clusters showing transient activity of GFP ranged from 2 to 52%. High levels of transient GFP activity and stable transformed callus lines were obtained with plasmid pTO134 containing a gfp gene with an enhanced CaMV 35S promoter and a bar gene with a 35S promoter in combination with Agrobacterium strain AGL0. These results suggest that the important variables in Agrobacterium-mediated transformation of wheat cells include media composition, Agrobacterium strain, plasmid vector and the addition of virulence-inducing agents such as acetosyringone. The conditions deemed optimal for transformation of wheat suspension cell lines were applied to scutella isolated from immature embryos and scutella-derived calli. Transient GFP expression in these tissues ranged from 10 to 75% and, while quite variable among and within cultivars, stably transformed scutellum-derived callus was obtained. Further studies with scutellum-derived calli suggested that variables such as duration of pre-inoculation culture and co-cultivation, as well as co-cultivation temperature, were also important. Optimisation of these variables resulted in the recovery of transformed wheat plants at a transformation frequency of 1.8%, which is comparable with other reports.

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