Additional phosphate stabilises uninterrupted growth of a Dioscorea deltoidea cell culture.

Suspension cells of Dioscorea deltoidea Wall (strain D-1) were maintained in a semicontinuous culture (SCC) in shake flasks at a high growth rate. It was shown that continuous propagation growth of this culture is unstable on Murashige's and Skoog's (MS) medium due to P starvation. On a P-enriched MS-medium the SCC was stable even at mean specific growth rates >0.3 day(-1). Highest volumetric concentrations of furostanol glycosides were obtained, when a P-enriched SCC was not further subcultivated but fed with sucrose. The investigated culture is able to control phosphate uptake and to prevent toxicity on media with excess P. High concentrations of cellular P(i) did not effect the ratio of furostanol to starch.

The boron requirement and cell wall properties of growing and stationary suspension-cultured chenopodium album L. cells

Suspension-cultured Chenopodium album L. cells are capable of continuous, long-term growth on a boron-deficient medium. Compared with cultures grown with boron, these cultures contained more enlarged and detached cells, had increased turbidity due to the rupture of a small number of cells, and contained cells with an increased cell wall pore size. These characteristics were reversed by the addition of boric acid (>/=7 &mgr;M) to the boron-deficient cells. C. album cells grown in the presence of 100 &mgr;M boric acid entered the stationary phase when they were not subcultured, and remained viable for at least 3 weeks. The transition from the growth phase to the stationary phase was accompanied by a decrease in the wall pore size. Cells grown without boric acid or with 7 &mgr;M boric acid were not able to reduce their wall pore size at the transition to the stationary phase. These cells could not be kept viable in the stationary phase, because they continued to expand and died as a result of wall rupture. The addition of 100 &mgr;M boric acid prevented wall rupture and the wall pore size was reduced to normal values. We conclude that boron is required to maintain the normal pore structure of the wall matrix and to mechanically stabilize the wall at growth termination.

[Separation of recombinant proteins by chromatography on vesicular sorbents]. / Razdelenie rekombinantnykh belkov s pomoshch'iu khromatografii na vezikuliarnykh sorbentov.

Vesicle chromatography, a recently developed method for separation of biomolecules, uses the vesicular packing (VP) material (clusters of microcapsules derived from plant cells), which was tested with respect to its application for the recombinant protein separation. Since VP has a well-defined separation limit, biomolecules are distributed in two separate peaks: large molecules are excluded and small molecules permeate through cell walls into the empty cell lumen. Recombinant proteins frequently form oligomers, which differ from monomers not only in size but also chemically and biologically. In the present study, separations of the recombinant proinsulin fusion protein oligomer and monomer, the recombinant human gamma-interferon monomer and dimer and recombinant tumour necrosis factor-alpha were investigated. For peak identification, the fractions and starting samples of the recombinant proteins were analysed by HPLC. The separations occurred without any sorption effects and with high efficiency and resolution of the protein peaks at a short column (10 cm). The VP is characterised by a high load ability, which favours the scale-up purification of the recombinant proteins. The combination of VP and HPLC is a considerable advance in biotechnology separation.

A parenchymatic medium solidifier for plant in vitro culture.

A new material for the solidification of liquid culture media was prepared from plant parenchyma tissues by mechanical subdivision, solute extration and dessication from ethanol. It is suitable for in vitro culture and propagation of callus as well as shoot tip cultures. The following plant materials have been grown by means of the new medium solidifier: shoot cultures of Betula pendula Roth, Gerbera jamesonii H. Bolus ex Hook and Floribunda rose "Triumph", callus tissues of Daucus carota L. and Chenopodium album L. The new solidifying material has special advantages over agar for application in the rooting phase of in vitro propagation.