Photoautotrophic cell lines

Selection procedures for isolation of photoautotrophic cell lines have been reviewed by Yamada and Sato (68). It should be noted that the culture of photoautotrophic cells has the potential for increasing secondary product synthesis. In instances where the synthesis of a particular secondary metabolite is regulated by the level of cellular differentiation, it is anticipated that the culture of photoautotrophic cells will have significant effects on production. Indeed, some alkaloids (69) vitamins (20). and components of volatile essential oils (70) have been produced by cultured green cells. 

Photoautotrophy may be introduced into a producing cell line by either of two methods (a) producing cell lines can be selected for photoautotrophy, or (b) protoplasts from a photoautotrophic cell line can be fused with protoplasts of the producing cell line and selection for both characteristics performed on subsequent fusion products. 

A number of factors have been studied for their influence on nicotine production. Of these the negative effect of auxins, and in particular 2,4-D, on alkaloid production is worth mentioning (202-204,211,220, 225,226,229,255). In root cultures the addition of indoleacetic acid (lAA) also reduces alkaloid production (196). Light was reported to inhibit nicotine formation (50,255). In a green cell suspension, however, increased nicotine levels were found on illumination (229). Ikemeyer and Barz (243) reported that a photoautotrophic cell line of N. tabacum did not produce nicotine, whereas a heterotrophic cell line did accumulate this alkaloid. Elicitation with a preparation of the fungus Phytophthora megasperma did not affect the nicotine levels of these cell lines. Addition of organic acids to the medium resulted in increased alkaloid formation in callus cultures (up to 3.25%) (230). For a review of the various cultural factors which influence secondary metabolism, the reader is referred to Mantell and Smith (255). 

To derive a cell line of photoautotrophic hairy roots, successive cultures of the photomixotrophic hairy roots were performed under the conditions of plain air (0.03% C02) and C02-enriched air (3.0% C02), and the initial concentration of sucrose, CSI, as a carbon source was lowered in a stepwise manner, that is, CSI= 10 (1st stage for 0-14 days), 5.0 (2nd stage for 14-28 days), 2.5 (3rd stage for 28-42 days), and 0 kg nr3 (4th stage for 42-56 days). 

Thus far, most photoautotrophic plant cells have been induced in cultures of calli or suspension cells [21,35-38]. Concerning plant hairy roots, Flores et al. [34] obtained the cell lines of photoautotrophic hairy roots of A. oppositifolia and D. innoxia through acclimation cultures with changes in sugar concentration in the medium and C02 tension in gas phase. However, detailed examinations have not been made to clarify the effects of culture conditions on the growth and photosynthetic activity of those photoautotrophic hairy roots. 

These suggest that the proliferation of the derived cell line of photoautotrophic pak-bung hairy roots is dependent entirely on photosynthesis for acquiring carbon and energy sources because DCMU blocks the reaction in photosynthetic process as shown in Fig. 6. In addition,based on the sensitive response to DCMU, a kind of photosynthesis inhibitor, the photoautotrophic hairy roots maybe used to detect some herbicides existing in the surroundings. 

Fig. 9. (left) Room-temperature EPR spectra of photoautotrophically grown cells of Synechocystis [(A) wild type and (B) D2-160(Y->F) mutant]. Heavy solid-line spectra were recorded immediately after illumination thin dashed-line spectra recorded 14 min after illumination. Figure source Debus, Barry, Babcock and McIntosh (1988) Site-directed mutagenesis identifies a tyrosine radical Involved in the photosynthetic oxygen-evolving system. Proc Nat Acad Sci, USA 85 429. 

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