In vitro propagation, regeneration, attempted tetraploid induction, and agrobacterium-mediated transformation of Euphorbia pulchurrima 'Winter Rose'(TM)

Poinsettia, Euphorbia pulchurrima, is the number one potted flowering plant in the United States. 'Winter Rose'(TM) is a popular cultivar with over one million plants sold each year. This research is aimed at establishing in vitro systems for application of biotechnology to poinsettia genetic improvement.; A protocol was established for in vitro axillary bud proliferation using greenhouse grown terminal buds. Buds were placed on Murashige-Skoog (MS) medium supplemented with benzlyaminopurine (BA). Explants produced the most axillary buds on medium containing between 2.2--8.8 muM BA. An organogenesis system was established using in vitro grown leaf tissues. The greatest amount of callus and shoots were produced from leaf midvein sections on MS medium containing 8.8--13.3 muM benzylaminopurine (BA) and 17.1 muM indole-3-acetic acid (IAA) for one month, and transferred to medium containing BA. Explants were also outplanted.; The effects of colchicine and oryzalin on callus and adventitious shoot formation and their ability to induce tetraploid formation of Euphorbia pulchurrima 'Winter Rose'(TM) was evaluated. Liquid or solid medium supplemented with colchicine or oryzalin was tested as well as a range of duration times between 1--4 days. Colchicine was less damaging to tissues at concentrations of 0.25 muM or 250.4 muM. Oryzalin inhibited regeneration from leaf tissues at all concentrations tested, and caused necrosis. A protocol was established for using flow cytometry to determine the ploidy level in poinsettia. Calluses and regenerated shoots from colchicine and oryzalin treatments were evaluated using the flow cytometer and found to be diploid. Colchicine can be considered a better tetraploid formation chemical for poinsettia due to callus and shoot regeneration.; The factors influencing Agrobacterium-mediated transformation of E. pulchurrima 'Winter Rose'(TM) were also evaluated. Kanamycin at 50 mg/L inhibited callus and shoot formation, and is a suitable selectable antibiotic for transformed cells in poinsettia. Variables evaluated included plasmid type (pBIl21, pMON690), the antibiotics CCK (cefotaxime, carbenicillin, and kanamycin) and acetosyringone. Since all tissues infected with agrobacterium (co-cultivation) died in 1--2 months, it appeared that poinsettia is highly sensitive to agrobacterium. Because of this premature death of infected tissues, other variables such as acetosyringone, and CCK could not be evaluated.