Transplantation of isolated nuclei into plant protoplasts : A novel technique for introducing foreign DNA into plant...

The uptake of isolated nuclei from Vicia hajastana Grossh. cells into protoplasts of an auxotrophic cell line of Datura innoxia P. Mill. was induced under the influence of polyethylene glycol and Ca(2+) at pH 6.8. The frequency of nuclear uptake varied from 0.8 to 2.3% and that of the recovery of prototrophic clones from 10(-5) to 6·10(-4). The prototrophic nuclear fusion products following nuclear uptake could be rescued by initial culture of the protoplasts in non-selective conditions and by the subsequent use of feeder cell layers to support the growth of surviving colonies on a selective medium. The presence of Vicia genomic DNA in some prototrophic clones was confirmed by dot-blot hybridization using Datura and Vicia DNA probes. In certain transformed clones, the recovery of prototrophy was accompanied by the restoration of morphogenetic potential. Welldeveloped shoots typical of wild-type Datura could be regenerated employing an appropriate regeneration medium.