Development and characterisation of an inducible system of betalain synthesis in cell cultures of beetroot(beta vulgaris)

The subject of this thesis was the development and characterisation of an inducible system for the production of betalains by red beet cultures. This was to be used as a model system to investigate the control of secondary metabolism in tissue culture. White lines of callus cultures of red beet, grown on solidified PGo salt cocktail medium containing 3% sucrose, 0.1mg/dm3 2,4-D, 0.5mg/dm3 kinetin were subjected to different light quantities and qualities, temperature and nutrient variations. In addition the effect of precursors was examined, feeding shikimic acid, tyrosine and DOPA in an attempt to see whether or not the control point could be detected from the biosynthetic pathway. It was also hoped to feed betalamic acid and cycloDOPA glucoside. However this necessitated the purification of standards in sufficient quantities so as to obtain these intermediates that did not exist commercially. This involved investigating different liquid chromatographic supports; Sephadex G-15, Dowex H+ and cellulose(20p). HPLC was also examined as a method to obtain standards. The results demonstrate the inducing effect of light and temperature. However this induction was characterised by a heterogeneous appearance. This result was considered best for the study of a heterogeneous response but not for the study of control of secondary metabolism. The feeding of precursors revealed that control was either further down the pathway at the intermediate betalamic acid or that each step of the biosynthetic pathway is turned on in concert in response to a signal