Explant Orientation and Polarity Determine the Morphogenic Response of Epicotyl Segments of Troyer Citrange

The morphogenic pathway of adventitious bud and shoot regenerationat the ends of Troyer citrange epicotyl cuttings is determinedby polarity and explant orientation. In explants planted verticallywith the basal end inserted in the medium, bud formation atthe apical end occurs by direct organogenesis. Bud growth andsubsequent shoot formation is markedly increased by the additionof 6-benzyladenine (BA) to the medium. This growth regulatoralso increases the number of buds formed. When they come intocontact with the culture medium, both the apical end and thebasal end of the cuttings form a vigorous callus with many xyllaryelements, more numerous in the calli from the basal end. Inthese calli, buds differentiate by a process of indirect organogenesis.This indirect regeneration pathway requires the addition of6-benzyladenine to the medium, and the number of buds formedis higher at the apical end than at the basal end of the cuttings.This pathway of regeneration is reduced as the position of thecuttings during incubation deviates from the normal uprightvertical position. Thus, for the basal end of the cuttings,the number of buds and shoots formed is higher when the explantsare placed vertically than when they lie on the surface of themedium. For the apical end, this number is higher in explantsplaced horizontally than when inserted vertically in the mediumin an inverted position. Copyright 1999 Annals of Botany Company Troyer citrange, Citrus sinensis x Poncirus trifoliata, explant orientation, histology, hormone dependence, morphogenesis, organogenesis, polarity, xylogenesis.     

Direct and Indirect Shoot Organogenic Pathways in Epicotyl Cuttings of Troyer Citrange Differ in Hormone Requirements and in their Response to Light

Bud differentiation by direct organogenesis at the apical endof Troyer citrange (Citrus sinensis[L]. OsbeckxPoncirus trifoliata[L].Raf.) epicotyl cuttings inserted vertically in a semi-solidculture medium did not require hormone additions. The numberof buds regenerated was slightly, but significantly, increasedwhen the incubation was performed in the light as compared tothe dark, and by the addition of benzyladenine (BA; 2.2 to 22µM) to the medium. Bud sprouting and subsequent shootformation required the addition of BA and was increased by lightto a higher extent than bud formation. The best response wasobtained with the highest BA concentration tested (22 µM).Regeneration through the indirect organogenic pathway at thetwo edges of the epicotyl cuttings when in contact with theculture medium did not occur in the absence of benzyladenine,which was an absolute requirement for callus development. Thebest regeneration response was obtained when the explants wereincubated in the light in the presence of 4.4 µM BA andan auxin. Indole-3-acetic acid (IAA; 5.8 µM) was moreeffective in increasing shoot formation than naphthaleneaceticacid (NAA; 0.54 µM). Higher NAA concentrations inhibitedshoot formation. Incubation in the dark or increasing the BAconcentration (22 µM) increased markedly callus growth,but inhibited both bud differentiation and sprouting, almostcompletely suppressing shoot formation. The conditions duringregeneration affected the rooting of the regenerated shoots.Rooting of 86% of the shoots was achieved in a medium with 2.7µM NAA and 2.6 µM indole-3-butyric acid. All therooted explants acclimated and survived transplanting. Underthe optimal conditions tested, the proliferation rate obtainedthrough the indirect regeneration pathway ranged from 60 to86 plants per seedling. Copyright 2000 Annals of Botany Company Troyer citrange, Citrus sinensisxPoncirus trifoliata, auxins, benzyladenine, direct organogenesis, hormone requirement, indirect organogenesis, light, morphogenesis, rooting.