In vitro propagation of cedar (Cedrela odorata L.) from juvenile shoots

Garriga, M (Garriga, Miguel); Caligari, PDS (Caligari, Peter D. S.). Univ Talca, Inst Biol Vegetal & Biotecnol, Talca, ChileCedrela odorata L. is one of the most important timber species currently traded in the Caribbean and Central America; however, it has been intensively exploited. In vitro techniques and clonal propagation can help to develop new plantations and assist in establishing improvement programs for this species. The aim of this study was to develop a protocol to establish in vitro conditions and to micropropagate this species from nodal explants from juvenile cuttings taken from field trees. Disinfection of node explants with 5% propiconazole CE 25 during 3 min resulted in 100% explant disinfection and 60% morphogenic response on those established explants. Shoot development was optimized by cultivating in vitro node explants in Murashige and Skoog basal medium supplemented with 2 mg L(-1) 6-bencilaminopurine and 3 mg L(-1) naphthaleneacetic acid. This medium resulted in 100% shoot development from the in vitro node explants with a 3.93 cm mean height. Rooting was also stimulated 6 wk after individualization of the regenerated plants on the same micropropagation medium with a mean of 3.9 roots per plant. In vitro plants did not show morphologic differences when compared to ex vitro seeds

Enhanced protoplast division by encapsulation in droplets: An advance towards somatic hybridisation in recalcitrant white lupin

Caligari, P.D.S. Instituto de Biologia Vegetal y Biotecnologia, Universidad de Talca, 2 Norte 685, Talca, ChileLupins are highly nutritious fodder and pulse crops but the greatest challenge in their genetic enhancement is the difficulty in obtaining hybrids through conventional sexual approaches. To bypass this, a procedure for the culture of hitherto recalcitrant lupin protoplasts is now being developed so that the somatic hybrids can be regenerated. This study provides a basis for a regime to culture lupin protoplasts. Cotyledonary protoplasts of white lupin (Lupinus albus) were plated in two diverse media for the evaluation of various plating regimes. The protoplasts divided in agarose as well as in Gelrite (TM) but embedding in agarose at 6 g L-1 concentration resulted in a higher rate of mitosis. Sodium alginate embedding inhibited protoplast division. Protoplast plating in the form of liquid suspension was significantly inferior to embedding. A filter paper substratum was clearly noxious to protoplast division. Vis-a-vis other designs of plating, a 400% improvement in protoplast elongation and division was achieved by plating in the form of 25 mu L droplets at the base of 60 mm x 15 mm Nunclon (TM) dish and overlaying with liquid medium. Better results in terms of protoplast elongation and division were obtained with K8p medium as compared to the AS medium. This report on lupin protoplast culture represents a significant breakthrough in the genus in which morphogenesis has not been described to date

Selection methods in plant breeding.

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Aspects of isolation underpinning mitotic behaviour in lupin protoplasts

This study reports on the influence of critical isolation factors on the subsequent culture of protoplasts of Lupinus albus L. Protoplasts were isolated from in vitro seedling cotyledons of five early maturing accessions in which protoplast yields and division frequencies appeared to be correlated as a high protoplast yield corresponded with a high division frequency. The overall difference among the accessions for mitosis was non- significant, although the highest yield and division frequency were observed in accession LA132, with Alban giving a significantly lower level. Accession Lucrop produced the lowest number of protoplasts, all of which collapsed during culture. Of the enzyme types used for tissue maceration, Pectolyase Y23, was significantly inferior to Macerase in terms of giving way to mitosis. The extent of division in Macerase- isolated protoplast population was 266% higher than that in the Pectolyase Y23- isolated one. The physiological maturity level of the explant, expressed in terms of developmental age, was optimal when 14 - 18- day- old seedling cotyledons were used for protoplast production and culture, rather than more mature ones, despite higher protoplast yields in the latter. On K8p medium, the protoplast division frequency was 129% greater when 18- day- old seedling cotyledons were used, than that with any other treatment. This work on protoplast culture of the potentially important lupin species, which is a pulse rich in dietary protein, oil and fibre, allows a further understanding of the biology, with an aim to advance lupin biotechnology

The chromosomal assessment of salt tolerant substituted tritipyrum using genomic fluorescent in situ hybridiization (FISH)

Wheat, although moderately tolerant to salt, can not be cultivated in many areas. However, in the triticeae tribe, some of the wild wheat relatives are highly tolerant, e.g. Thinopyrum bessarabicum, which grows on the sea shore. Eight primary hexaploid tritipyrum lines, amphiploids between Triticum durum and Thinopyrum bessarabicum have been produced which can set seed in at least 250 mM NaCl. These tritipyrums (2n=6x=42, AABBEbEb) due to reasons such as brittle rachis, continuous production of tillers, late maturity, tall stature and meiotic instability will not fulfill the requirements of a successful commercial salt tolerant crop. To overcome such problems the substituted tritipyrum, in which selected Eb chromosomes are replaced by D genome chromosomes of 6x wheat, was produced from 6x tritipyrum x 6x wheat hybrids (F1: 2n=6x=42, AABBDEb) followed by selfing and backcrossing with 6x tritipyrum. The fertile plants among the above progenies were screened by the genomic fluorescent in situ hybridization technique to identify their Eb and D chromosome constitution. This study showed that producing tritiprum with variable numbers of Eb and D genome chromosomes is feasible and that FISH is a useful technique for determining the number of Eb chromosomes present