During the first 8-12 days of cultivation in a nutrient solution containing IAA, inositol and kinetin freshly isolated carrot root explants develop into a chlorophyllous and photosynthetically active tissue culture. Electron microscopy, low temperature pigment absorption spectra and fluorescence induction profile recording as well as the determination of the activity of some enzymes (ribulosebisphosphate carboxylase, phosphoenolpyruvate carboxylase) and CO2-fixation experiments were carried out. Based on the results, a sequence of developmental stages of the photosynthetic system will be proposed. During the autotropic period from about the 20th day of culture onward the light reaction system of these tissue cultures is quite comparable to that of carrot leaves; however, some differences in the CO2-fixation mechanism were observed.
Bender, L., Joy IV, R. W. and Thorpe, T. A. 1987. Studies on [14C]‐glucose metabolism during shoot bud induction in cultured cotyledon explants of Pinus radiala.Excised cotyledons of Pinus radiata D. Don, cultured under shoot‐forming (plus N6‐benzyladenine) and elongating (minus N6‐benzyladenine) conditions, were fed U‐[14C]‐glucose for 3 h in the light followed by a 3 h chase period immediately after excision (day 0) and after 3 days of culture (day 3). The incorporation of l4C into individual soluble metabolites as well as into protein was followed. No labelled citrate could be detected at day 0, however, a flow of 14C from glucose to glutamate/ glutamine occurred. During this stage the synthesis of glutamine strongly increased in the cotyledons supplied with N6‐benzyladenine, which suggests a positive influence of this cytokinin on nitrogen incorporation prior to differentiation. After 3 days of cultivation large amounts of labelled citrate were detected. An increased incorporation of label into protein due to the cytokinin treatment was not detected during the early culture period (days 0 and 3). Labelled amino acids were incorporated into protein to different degrees, but this was not influenced by the hormonal treatment.
Chromoplasts, which exist in the ceils of freshly isolated carrot root explants, seemed to be transformed in thylakoid containing plastids, and chlorophyll formation was initiated if the explants were cultured in a liquid medium containing inositol and IAA as a hormonal supplement. This process was intensified when kinetin was also added, but no dependence on a sucrose supply could be found.A sucrose supply of 2% in conjunction with the combination of all three hormones, however, was needed to achieve maximal thylakoid formation including stacking in individual chloroplasts and for the very extensive chloroplast multiplication in explants growing with high cell division activity. It should be noted that the number of plastids per cell is strongly increased by the sucrose supplement which leads also to starch accumulation. However, no transformation into chloroplasts occurred without the hormonal stimulus.
CO2 fixation experiments with photosynthetically active 3-week-old carrot callus cultures (60–90 μg chlorophyll∙g fresh weight−1) were carried out in light and darkness in nutrient solution containing NaH14CO3. Samples were taken and analyzed for labelled metabolites after 15 s and 4 min of 14CO2 fixation. The results show that in the light both the Calvin cycle and phosphoenolpyruvate carboxylation were observable, whereas in the darkness CO2 was fixed only by phosphoenolpyruvate carboxylation. However, extractable phosphoenolpyruvate carboxylation activity was reduced by about 80% upon darkening the light-grown cultures for 1.5 h.
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