Transformation and regeneration of
View Patent ↗The invention relates to a novel transformation method for plants of the genus Allium , in particular to onion plants. Plants transformed by the method are also provided. The method preferably involves an Agrobacterium tumefaciens -mediated transformation, more preferably involving immature embryos as the explant source and employing a binary vector.
1. A method of transforming an Allium cepa plant comprising the following steps:
(a) transforming embryo cells of the Allium cepa plant with DNA sequences via a vector or direct gene transfer to produce transformed plant material, wherein transformation is achieved by:
(i) wounding the embryo cells and transferring the embryo cells into a suspension of Agrobacterium,
(ii) transferring the embryo cells from step (i) to a culture medium;
(iii) co-cultivating the embryo cells and the Agrobacterium attached to the embryo cells for a period of 1–12 days from the end of step (ii);
(b) selecting the transformed plant material derived from step (a), by transferring the embryo cells to a selection medium containing the appropriate selection agents to kill the agrobacteria and preferentially grow the transgenic embryo cells to produce transformed plant material;
(c) culturing the transformed plant material from (b) to produce secondary embryos and regenerating transformed plant material from the secondary embryos; and
(d) obtaining a transformed Allium plant from the secondary embryo;
wherein the method of transforming is carried out without a passage through a callus phase.
2. A method of transforming an Allium cepa plant comprising the following steps:
(a) transforming embryo cells of the Allium cepa plant with DNA sequences via a vector or direct gene transfer to produce transformed plant material, wherein transformation is achieved by:
(i) wounding the embryo cells and transferring the embryo cells into a suspension of Agrobacterium containing a plasmid with a functional T-DNA region that is capable of transfer to plant cells,
(ii) transferring the embryo cells from step (i) to a culture medium;
(iii) co-cultivating the embryo cells and the Agrobacterium attached to the embryo cells for a period of 1–12 days from the end of step (ii);
(b) selecting the transformed plant material derived from step (a), by transferring the embryo cells to a selection medium containing the appropriate selection agents to kill the agrobacteria and preferentially grow the transgenic embryo cells to produce transformed plant material;
(c) culturing the transformed plant material from (b) to produce secondary embryos and regenerating transformed plant material from the secondary embryos; and
(d) obtaining a transformed Allium plant from the secondary embryo;
wherein the method of transforming is carried out without a passage through a callus phase.