Comparison of Starch and ADP-Glucose Pyrophosphorylase Levels in Nonembryogenic Cells and Developing Embryos from Induced Carrot Cultures

Abstract

Cultures of carrot (Daucus carota L.) in a medium without added 2,4-dichlorophenoxyacetic acid were separated into fractions of embryos at different stages of development (large globular and heart, torpedo, and germinating) and nonembryogenic cells. The average starch content per cell in these fractions was similar. However, due to the smaller sizes of the cells of the embryos relative to the nonembryogenic cells, starch content per weight of tissue was higher in the embryos. The ADP-glucose pyrophosphorylase activity per cell in the nonembryogenic cells was double that of the embryo cells. Furthermore, the ratio of ADP-glucose pyrophosphorylase to starch was over 2-fold higher in the nonembryogenic cells, indicating that starch content is not simply determined by ADPglucose pyrophosphorylase levels. ADP-glucose pyrophosphorylase activity of all culture fractions was directly proportional to the level of a single 50 kilodalton polypeptide detected by immunoblot analysis, using antiserum raised to the purified spinach leaf enzyme. In the same immunoblot analysis, novel polypeptides of 63 and 100 kilodalton were detected in embryos but were absent from nonembryogenic cells. This is one of the few reported examples of specific proteins which differentially accumulate in embryos and nonembryogenic cells.

Differential Accumulation of Biotin Enzymes during Carrot Somatic Embryogenesis

“…Cell (22). Transfer of such cultures to medium lacking 2,4-D resulted in the organized development of the embryogenic cell clusters into embryos, whereas the nonembryogenic cells continued to divide but did not undergo embryogenesis (10). Embryogenic cell clusters and developing embryos at specific stages of development were examined for biotin enzymes and the accumulation of biotin-containing polypeptides.…”

“…Embryos at specific developmental stages and nonembryogenic cells were obtained from (-)2,4-D cultures by Ficoll density gradient centrifugation and size fractionation (10). The resultant fractions were 80 to 95% pure as estimated by microscopic examinations.…”

“…Cell cultures were initiated from the secondary phloem of carrot roots (Daucus carota var Danver) and maintained essentially as previously described (10,22) …”

“…The somatic embryogenesis system of carrot (8) enabled the isolation of large quantities of embryos at specific stages of development (6,7,10,22). We report the changes in the activities of the four biotin enzymes of plants (acetyl-CoA carboxylase, propionyl-CoA carboxylase, 3-methylcrotonyl-CoA carboxylase, and pyruvate carboxylase) and the changes in the accumulation of the biotincontaining polypeptides during carrot somatic embryo development. …”

Effect of culture medium and light conditions on the morphological characteristics and carbohydrate contents of Medicago strasseri calli

“…Since carbohydrates serve as sources of carbon and energy in callus formation (Rawal et al 1984, Swarnkar et al 1986) and since the characteristics of calli are largely determined by their carbohydrate composition (Keller et al 1988, Naidu and Kishor 1995, Vu et al 1995, we studied these compounds in the different calli formed to determine which medium/a and composition(s) are the most suitable for callus induction.…”