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Metabolic changes in tobacco plants grown in the presence of sucrose
Garčeková, Květa ; Ryšlavá, Helena (advisor) ; Folwarczna, Jitka (referee)
Metabolism of plants grown in vitro is affected by the composition of culture medium. Beside essential nutrients culture medium are often composed of sucrose as well. The aim of this study was to find out, if additional source of carbon-sucrose affected activity of phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31), NADP-malic enzyme (NADP-ME, EC 1.1.1.40), and pyruvate, phosphate dikinase (PPDK, EC 2.7.9.1). Activities of these enzymes were determined in different parts of tobacco plants (Nicotiana tabacum L., cv. Petit Havana SR1) grown in vitro in culture medium: Murashige-Skoog agar containing sucrose (SR1+S) and in plants grown without sucrose (SR1-S). Fresh weight activity and specific activity of studied enzymes in SR1+S plants were increased. In upper leaves fresh weight activity of PEPC, NADP-ME and PPDK was increased 1.7- times, 1.4-times and 1.9-times, respectively. Also the protein content was higher in SR1+S plants. The results indicate that metabolic pathways catalyzed by these enzymes are in the presence of sucrose more involved. Key words: Nicotiana tabacum, L., NADP-ME, PEPC, PPDK, sucrose
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Production of heterologous proteins in plants - human papillomavirus (HPV 16) derived antigens
Folwarczna, Jitka ; Čeřovská, Noemi (advisor) ; Ryšánek, Pavel (referee) ; Kumar, Jiban (referee)
5 Abstract Even though prophylactic vaccine against human papillomavirus (HPV) is currently licensed, infections by the virus continue to be the major health problem mainly in developing countries. Considerable effort is being devoted to preparation of therapeutic vaccine and to decrease of the production costs of current vaccine. Viral proteins such as the E7 oncoprotein and the L2 capsid protein from HPV type 16 are promising targets for the development of the experimental anti-HPV vaccine. The aim of our work was optimization of expression of mutagenized E7 oncoprotein (E7ggg) fused to the C-terminus of Tobacco mosaic virus (TMV) coat protein (CP) or Potato virus X (PVX) CP in viral vectors derived from these plant viruses. The impact of linkers connecting CP and E7ggg fusion partners on expression and stability of fusion proteins was examined. The fusion proteins were first expressed in Escherichia coli (E. coli) MC1061 to assess the characteristics of the recombinant protein prior to their transient expression in both non-transgenic or transgenic Nicotiana benthamiana (N. benthamiana). We have obtained the high level expression in E. coli, but most of the expressed proteins based on TMV CP remained in insoluble inclusion bodies. To increase the ratio of soluble protein various molecular...
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Metabolic changes in tobacco plants grown in the presence of sucrose
Garčeková, Květa ; Ryšlavá, Helena (advisor) ; Folwarczna, Jitka (referee)
Metabolism of plants grown in vitro is affected by the composition of culture medium. Beside essential nutrients culture medium are often composed of sucrose as well. The aim of this study was to find out, if additional source of carbon-sucrose affected activity of phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31), NADP-malic enzyme (NADP-ME, EC 1.1.1.40), and pyruvate, phosphate dikinase (PPDK, EC 2.7.9.1). Activities of these enzymes were determined in different parts of tobacco plants (Nicotiana tabacum L., cv. Petit Havana SR1) grown in vitro in culture medium: Murashige-Skoog agar containing sucrose (SR1+S) and in plants grown without sucrose (SR1-S). Fresh weight activity and specific activity of studied enzymes in SR1+S plants were increased. In upper leaves fresh weight activity of PEPC, NADP-ME and PPDK was increased 1.7- times, 1.4-times and 1.9-times, respectively. Also the protein content was higher in SR1+S plants. The results indicate that metabolic pathways catalyzed by these enzymes are in the presence of sucrose more involved. Key words: Nicotiana tabacum, L., NADP-ME, PEPC, PPDK, sucrose
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