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Background  Impatiens ( Impatiens walleriana ) is a top selling floriculture crop. The potential for genetic transformation of Impatiens to introduce novel flower colors or virus resistance has been limited by its general recalcitrance to tissue culture and transformation manipulations. We have established a regeneration and transformation system for Impatiens that provides new alternatives to genetic improvement of this crop.    Results  In a first step towards the development of transgenic INSV-resistant  Impatiens , we developed an efficient plant regeneration system using hypocotyl segments containing cotyledonary nodes as explants. With this regeneration system, 80% of explants produced an average of 32.3 elongated shoots per initial explant plated, with up to 167 elongated shoots produced per explant. Rooting efficiency was high, and 100% of shoots produced roots within 12 days under optimal conditions, allowing plant regeneration within approximately 8 weeks. Using this regeneration system, we developed an efficient  Agrobacterium -mediated  Impatiens  transformation method using  in vitro  multiple bud cultures as explants and a binary plasmid (pHB2892) bearing  gfp  and  nptII  genes. Transgenic  Impatiens  plants, with a frequency up to 58.9%, were obtained within 12 to 16 weeks from inoculation to transfer of transgenic plants to soil. Transgenic plants were confirmed by Southern blot, phenotypic assays and T 1  segregation analysis. Transgene expression was observed in leaves, stems, roots, flowers, and fruit. The transgenic plants were fertile and phenotypically normal.    Conclusion  We report the development of a simple and efficient  Agrobacterium -mediated transformation system for  Impatiens . To the best of our knowledge, there have been no reports of  Agrobacterium -mediated transformation of Impatiens with experimental evidence of stable integration of T-DNA and of  Agrobacterium -mediated transformation method for plants using  in vitro  maintained multiple bud cultures as explants. This transformation system has the advantages of 1) efficient, simple and rapid regeneration and transformation (with no need for sterilization or a greenhouse to grow stock plants), 2) flexibility (available all the time) for  in vitro  manipulation, 3) uniform and desirable green tissue explants for both nuclear and plastid transformation using  Agrobacterium -mediated and biolistics methods, 4) no somaclonal variation and 5) resolution of necrosis of  Agrobacterium -inoculated tissues.

bmc plant biology

Development of Efficient Plant Regeneration and Transformation System for Impatiens Using Agrobacterium tumefaciens and Multiple Bud Cultures as Explants