Manuel L. Robert

In vitro propagation of Agave fourcroydes Lem. (Henequen)

In vitro plant regeneration of Agave fourcroydes Lem. (Agavaceae) is described. Results suggest that the NO3-:NH4+ balance in the culture medium is a key factor controlling callus growth and organogenesis in rhizome cultures. Stem callus showed limited organogenic capacity, but high cytokinin concentrations induced adventitious shoot formation on stem explants. When these shoots were excised and subcultured, new callus formed at their base from which new shoots arose. The shoots from stem explants and rhizome callus formed extensive root systems in vitro and were transferred to pot culture with a 90% survival rate.

Glutamate dehydrogenase activity in normal and vitrified plants of Agave tequilana Weber propagated in vitro.

Agave tequilana stem explants were used to produce adventitious shoots under a set of different water potentials induced by different concentrations of gelrite in the medium. At high water potentials all shoots were vitrified; as the medium water potential became more negative the degree of vitrification decreased but the number of shoots per explant also diminished. The enzymes NADH and NAD-GDH (EC. 1.4.1.2) were measured along the water potential gradient. GDH activity was high in the non-vitrified tissues and decreased significantly in the vitrified ones.

Growth promoting activity of certain penicillins on cultivated cells of Bouvardia ternifolia

Abstract A growth promoting effect similar to that of 2,4-D is induced in Bouvardia ternifolia calli and cell suspension cultures by some beta lactam antibiotics but not by others. It was found that this effect was exerted only by those penicillins that contain an auxin-like moiety associated with the 6-aminopenicillanic acid, such as carbenicillin or penicillin G. Phenylacetic acid and 6-aminopenicillanic acid were inactive on their own.

Nitrogen Metabolism in Canavalia ensiformis (L.) DC.: II. Changing Activities of Nitrogen-Assimilating Enzymes during Growth

Summary The ontogeny of glutamine synthetase (EC 6.3.1.2), glutamate synthase (EC 1.4.1.14), asparagine synthetase (EC 6.4.5.4) and glutamate dehydrogenase (EC 1.4.1.2) has been followed in cotyledons, roots, stems and leaves of young plants of Canavalia ensiformis . The four enzymes were found in all regions. Glutamine synthetase activity increased in the leaves with time and remained low in the cotyledons. Initially glutamate synthase activity in the cotyledons was NADH-dependent but later became Fd-dependent. Asparagine synthetase specific activity was highest in the roots until day 5 after which activity was similar in all regions. While glutamate dehydrogenase specific activity decreased with time in the leaves it increased in the cotyledons and roots. We suggest that in the early stages of seedling growth asparagine synthetase and glutamate dehydrogenase both play a fundamental role in the reassimilation of ammonium released from canavanine. When the plant acquires full photosynthetic capacity the glutamine synthetase/glutamate synthase pathway becomes the principal pathway of reassimilation in the aerial parts of the plant.

Nitrogen Metabolism in Canavalia ensiformis L. DC.: I. Arginase and Urease Ontogeny

Summary The ontogeny of arginase and urease has been followed in cotyledons, roots, stems and leaves of young Canavalia ensiformis plants. Both enzymes were found in all organs. Arginase activities were highest in stems and roots, whereas urease activity was highest in the cotyledons. Urea and ammonia pools were measured in all organs studied and, in general, decreased with time, except in leaves where a marked increase in ammonia was observed. The data are discussed in relation to the mobilization and distribution of canavanine from cotyledons to other regions during germination and subsequent seedling growth.

Liquid In Vitro Culture for the Propagation of Arundo donax

We describe a simple and inexpensive plant micropropagation system for giant reed (Arundo donax L.) that uses axillary buds from the lateral stems of elite plants selected from field- or nursery-grown plants. The buds, attached to the stems are cultured in stationary liquid MS culture medium, supplemented with indole 3-acetic acid and kinetin. This formulation is the only one required for all the stages. Contrary to what happens in semisolid medium where roots are not formed, the plants cultured in liquid medium are whole plants with shoots and roots that develop at the same time. The survival rate of these plants when transferred to soil is close to 100% during acclimatization. A clonal line of 900 plants from a single mother plant can be produced in 4 months.

Tissue Culture Methods for the Clonal Propagation and Genetic Improvement of Spanish Red Cedar ( Cedrela odorata )

The choice of a method to culture red cedar tissues depends on the final objectives pursued. If homogeneous clonal material is required for experimental purposes, the easiest way is to generate the lines through adventitious shoot induction from seedlings germinated from seeds. If the objective is to generate high yielding material for plantation purposes, the choice will be the same method but starting from mature vegetative tissues from selected elite plants. Most of the process are the same, but the initial steps are less efficient and much more elaborate. If the purpose is to generate lines with new genetic characteristics through somaclonal variation, mutagenesis, or genetic transformation, somatic embryogenesis will be required. No single method in its present form is suitable for all purposes. Eventually, the efficient production of somatic embryos from rejuvenated shoots collected from mature selected plants is the ideal way to culture this species, but for the time being we have to choose one or the other. In this chapter, we present a grafting procedure to rejuvenate and maintain mother plants in the greenhouse and the in vitro culture systems we have developed for the production of Cedrela odorata propagules using explants from both young seedlings and mature tissues from selected old trees. Using a modified TY17 medium and the BioMINT(®) temporary immersion system, we obtained high multiplication and ex vitro transplantation rates for efficient large-scale propagation of this species.

Somatic Embryogenesis in Agave: An Overview

This chapter describes the details of somatic embryogenesis of some species of the genus Agave. Various factors that influence the induction of the embryogenic process, such as explant type, growth regulator type, growth regulator concentration and genotype, are discussed. Furthermore, embryo development and conversion, as well as the first works using temporary immersion bioreactors, are also discussed.

Somatic Embryogenesis in Temporary Immersion Bioreactors

Somatic embryogenesis is a very useful micropropagation technique, due to its high multiplicative capacity that offers the potential for large-scale propagation using temporary immersion bioreactors. The temporary immersion system is mainly based on the contact of plant tissue with culture medium by certain cycles of immersion, avoiding the problems of hyperhydricity, malformed embryos, and low conversion rates, which occur in continuous immersion systems. The automation of some or all of the phases of the process of somatic embryogenesis in a bioreactor could reduce labor and gellant costs and increase micropropagation efficiency, allowing high-quality plantlets to be obtained through more efficient and controlled protocols. This chapter describes the different types of temporary immersion bioreactors that have been used to increase or scale somatic embryogenesis in different plant species.