and cultivated in the greenhouse. The average number of flowers at anthesis per plant(Fig. 3B) was higher in the transgenic plants of clone 301 as a consequence of a moreprofuse flower bud production (Fig. 3A). The transgenic plants produced 30% moreflowers per plant than control (Fig. 3C). Moreover, transgenic plants retained their abilityto flower in June whereas control plants flowered sporadically and entered vegetativephase. In August only transgenic plants were still able to produce new flower buds (anaverage number of 7.9 buds per plant).4. DiscussionThe non-conventional breeding program based on genetic engineering wassuccessful in Osteospermum to modify ornamental traits by using rol genes ofAgrobacterium rhizogenes (Giovannini et al., 1999), and to obtain virus tolerant plantswith the N protein gene of tomato spotted wilt virus (Vaira et al., 2000). Preliminaryresults from this work show that constitutive expression of a single Arabidopsis floweringtime CO gene increases the production of flowers in the ornamental plant O. ecklonisduring the normal flowering period (February-May) and extends the period of floweringduring not permissive conditions (June to August). The early flowering phenotypereported over-expressing CO gene in Arabidopsis (Putterill et al., 1995) is not observed in35SCO Osteospermum plants of clone 301, probably because in this species the transitionfrom vegetative to reproductive phase is under the control of a different genetic pathway(e.g. vernalisation pathway). On the other hand the extension of flowering in thetransgenic plants could be related to the continuous expression of the CO gene and itsregulatory effect on other genes. It is our purpose to evaluate the behaviour of additionaltransgenic clones expressing CO gelne and extend the evaluation period along the year.The final aim is the understanding of the CO gene potential for the production of O.ecklonis genotypes that flower even in not permissive conditions.AcknowledgementsThe authors would like to thank Dr. George Coupland and Dr. Phil Mullineaux forproviding the CONSTANS gene and the pGREEN vector, respectively. We are alsograteful to Jemma Miller for the English revision of this work.ReferencesAllavena A., Giovannini A., Berio T., Spena A., Zottini M., Accotto G.P., and VairaA.M., 2000. Genetic engineering of Osteospermum SPP: a case story. Acta Hort.508:129-133.Giovannini A., Zottini M., Morreale G., Spena A., and Allavena A., 1999. Ornamentaltraits modification by rol genes in Osteospermum ecklonis transformed withAgrobacterium tumefaciens. In Vitro Cell. Dev. Biol.-Plant 35:70-75.Klimyuk V., Carrol B.J, Thommas C.M., and Jones GDJ, 1993, Alkali treatment for rapidpreparation of plant material for reliable PCR analysis, The Plant J.3: 493-494.Putteril J., Robson F., Lee K., and Coupland G., 1993, Chromosome walking with YACclones in Arabidopsis : isolation of 1700 kb of contiguous DNA on chromosome 5,including a 300kb region containing the flowering time gene CO. Mol.Gen.Genet. 239:145-157.Putteril J., Robson F., Lee K., Simon R., and Coupland G., 1995, The CONSTANS geneof Arabidopsis promotes flowering and encodes a protein showing similarities to zincfinger trancription factors, Cell 80: 847-857.Suárez-López P, Wheatley K., Robson F., Onouchi H., Valverde F., and Coupland G,2000, CONSTANS mediates between the circadian clock and the control of floweringin Arabidopsis, Nature 410:1116-1120.Vaira A.M., Berio T., Accotto G.P., Vecchiati M., and Allavena A., 2000. Evaluation of165 Source.


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