Growth, ageing and death of a photoautotrophic plant cell culture

Abstract

Batch cultures of photoautotrophic cell suspensions of Chenopodium rubrum L., growing in an inorganic medium on CO2 under a daily balanced light-dark regime of 16: 8 h could be maintained for approximately 100 d without subcultivation. The long-lived cultures showed an initial cell division phase of 4 weeks, followed by a stationary phase of another 4 weeks, after which ageing and progressive cell death reduced the number of living cells and the cultures usually expired after another 3-4 weeks. These developmental phases of the cell culture were characterised with respect to photosynthetic performance, dark respiration, content of phytohormones and capacity of cell division. Cell division of the majority of the cells finished in the G1- or G0-phase of the cell cycle, caused by a pronounced decline in the endogenous levels of auxin and cytokinins. Supply of these growth factors to resting cells resulted in resumption of cytokinesis, at least by some of the cells. However, responsiveness to the phytohormones declined during the stationary phase, and subcultivation was no longer possible beyond day 60 when the phases of ageing and death commenced. Ageing was characterised by a further decline in the photosynthetic capacity of the cells, by a climacteric enhancement of dark respiration, but also by a slight increase in the level of IAA and cytokinins concomitant with a decrease in ethylene. Similarities and differences between the development of batch-cultured photoautotrophic cells of C. rubrum and that of a leaf are discussed with respect to using the cell culture as a model for a leaf.

Purification of 3‐indolylacetic acid by solid phase extraction

“…Thus the use of both phosphoric [17] and acetic acid [19] for acidification of plant extract prior to SPE was reported. Elution of retained IAA was accomplished by using either absolute [17,19] or 80% methanol [18]. The use of 80% methanol containing 1% of acetic acid was also described [15].…”

“…Typically, the plant extract of IAA in 80% methanol is acidified only after it has been transformed into an aqueous phase by vacuum evaporation [18,19,23]. To eliminate this time-consuming step and replace it by a simple dilution procedure, we studied the influence of various methanol concentrations on IAA retention on the C18 matrix.…”

“…Irrespective of the solution used for extraction (which may be phosphate buffer of pH 6.5 [16,17], absolute methanol [18], 90% methanol [19], or 80% methanol [20]), such SPE is based on the retention of the protonated form of IAA on a non-polar C18 matrix. To achieve this, methanolic extracts are reduced to an aqueous phase by evaporation in vacuo and, similarly to phosphate buffer extracts, acidified to shift the acid-base balance of IAA in favour of its uncharged form.…”

“…To achieve this, methanolic extracts are reduced to an aqueous phase by evaporation in vacuo and, similarly to phosphate buffer extracts, acidified to shift the acid-base balance of IAA in favour of its uncharged form. Subsequent elution of retained IAA is usually performed by application of either absolute [17,19] or 80% methanol [18] or by application of 80% methanol containing 1% of acetic acid [15].…”

Torn Between Nature and Lab: A Dying Breed of Plant Scientists?

“…To characterize the individual developmental stages of that cell culture, we investigated the respective patterns and metabolic interconversions of these phytohormones and their derivatives (Fußeder et al 1988b). Bearing in mind the "stay-green" mutants (e.g., Hauck et al 1997) and the green islets in pathogen-attacked leaves (Faiss et al 1997) we identified the point of no return from ageing in our cell cultures as that state at which addition of phytohormones could not reverse the process of ageing (Peters et al 2000). One of the genes whose expression is under the control of auxin turned out as the gene of a glycan-exohydrolase OrGEH that is essential for growth of the cell wall and therefore also for the growth of the cell culture (Dominguez et al 2014).…”