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System with AC generator for cell electroporation
Lukoianov, Petr ; Folprecht, Martin (referee) ; Červinka, Dalibor (advisor)
This bachelor thesis is observing a system with AC generator for cell electroporation. The first paragraph keeps an eye on description of methodology of biophysics mechanisms. In the 2nd part several kinds of devices were described. In the last paragraph you can see a non-assembled construction and operating principle of the device. Apart from that the control circuit logic explored and device manual is included.
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Gene expression in chicken embryo: micromanipulation and visualization methods
Bendová, Michaela ; Dvořák, Michal (advisor) ; Hirsch, Ivan (referee) ; Krylov, Vladimír (referee)
The aim of this work was to obtain better insight into the principles of cell structures and organs in the chicken embryo development. To reach this goal special methods of micromanipulations and visualization in vitro, ex vivo, ex ovo and in ovo were implemented and adjusted. These methods were used to study gene expression in neural crest development and eye development. In the course of long term research in our laboratory we observed that oncoprotein v-Myb influences the development of the neural crest and has the capacity to change natural cell fate. We performed a series of experiments to investigate v-Myb protein influence on neural crest cells differentiation, especially melanocyte lineage development, and its influence on gene expression in the neural crest. Therefore we focused on Gremlin 2 (PRDC), the gene upregulated by v-Myb in the neural crest. The established procedure of electroporation in ovo was adjusted to transfect cells of the developing eye and used to study gene expression during lens induction. The results obtained from chicken embryo experiments endorsed the study performed on mouse embryos. Futhermore, the electroporation technique was slightly modified for manipulations of the neural retina in the developing eye in ovo. Thereafter, the retinas were processed ex vivo and...
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EKG synchonization of electroporation pulses
Nohel, Michal ; Mézl, Martin (referee) ; Novotná, Veronika (advisor)
Bachelor thesis is focused on cells electroporation principle and its types. It further describes thermal and nonthermal ablation techniques with an emphasis on irreversible electroporation. It contains an overview of fields, where medical procedures in combination with ECG signal are used, and an overview of techniques and measurements of the electrocardiographic signal. Further, the sensitivity of the human body to electroporation pulses is discussed. During electroporation, short high-voltage pulses are applied. Ventricular fibrillation may occur if the electroporation pulse is applied to the vulnerable period of the heart rhythm. Synchronization of pulse application with heart activity of the patient is necessary for health and safety reasons in clinical practice. The practical part of the thesis is about a LabVIEW program for this kind of synchronization. The created program is tested and the obtained results achieved are evaluated in terms of practical use.
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Regulation of gene expression in anaerobic parasitic protist and practical application of knowledge
Brzoň, Ondřej ; Zubáčová, Zuzana (advisor) ; Horváthová, Lenka (referee)
Because of its importance for every single cell, regulation of gene expression is often studied at all levels today. On the other hand, rather sparse amount of information is available for protists and our understanding of their gene expression is nearly limited to several model organisms. Three anaerobic human parasites have been studied in details: Trichomonas vaginalis, Entamoeba histolytica and Giardia intestinalis. Most recently, some others anaerobic protists (e.g. Entamoeba invadens or some species of Spironucleus genus) have been researched as well. This work is focused on the structure of promoter regions in protein coding genes and application of this knowledge in construction of transfection systems as one of the most effective and powerful tools in molecular and cell biology.
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Controlled high voltage source
Chloupek, Jiří ; Hanák, Pavel (referee) ; Koton, Jaroslav (advisor)
The aim of this master thesis is to design a high voltage source for the generation of bipolar pulses, which are suitable for the process of reversible and irreversible electroporation. Further design is electronic control. The first part of the thesis analyzes the theoretical knowledge about the process of electroporation, the second part describes the principles of realization of such sources, the next part deals with the design of the source and its control. The penultimate part is a description of the user manual and the last part is dedicated to the measurement.
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Transformace zelené řasy \kur{Desmodesmus quadricauda}
ZAHRADNÍKOVÁ, Jana
The aim of this study was to investigate if electroporation can be used for a preparation of stable transformants of the green alga Desmodesmus quadricauda. I tested a sensitivity of D. quadricauda cells to several antibiotics in order to choose a suitable selection marker. I also tried to optimize electroporation conditions. I tested several electroporation methods and I tried transformation of cells of asynchronous cultures and in different stages of the cell cycle in synchronous culture. I tested the effect of sonication and glass beads on cell wall of coenobia in order to slightly damage it, release individual cells and, thus, affect the transformation efficiency.
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Controlled source of pulsed electric field
Burian, Josef ; Kubánek, David (referee) ; Koton, Jaroslav (advisor)
The aim of this master thesis is to design a power source for generating pulsed electric field for the needs of the technological process of electroporation of fruit musts and mashes. To design further the network of switched capacitors and inductors based on the required pulse and to design and implement the basic control unit together with the switching transistors. The thesis will include calculations and simulations used in the draft, also design solutions and measured values. This thesis is divided into several basic parts. In the first part there is discussed in detail the theoretical knowledge of electroporation and the desired characteristics of generated pulses are chosen according to this knowledge. Each part of electroporation workplace is described in the second part of this thesis, beginning from the source through the control system to the electrode chamber. For each of these parts are given different possible alternatives. In the next chapter is already proceeded to the design of the source. There are listed required parameters of the pulses and according to them calculations and the design are gradually carried out. Another chapter deals with the simulations, which are used to verify the calculated values and conditions in the electroporated sample. Last but one part discusses the mechanical design of the workplace. There are described all problems of the construction and commissioning of the product. The last section is dedicated to the workplace measurement and analysis of the measured results.
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Transformation of \kur{Chlamydomonas reinhardtii} ptDNA
HUSÁKOVÁ, Jana
The aim of this master thesis was to test and compare two available methods of genetic transformation (biolistics, electroporation) of the plastid genome of green algae Chlamydomonas reinhardtii. For biolistic transformation a wide range of experimental parameters which generally influence ptDNA transformation efficiency was optimized: physiological condition of acceptor cells, type and size of microparticles, pressure of propulsion gas (helium), length of projectile trajectory, transformation of cells directly on selective medium (containing spectinomycin 150 ?g/ml) or on nonselective medium (without spectinomycin) and form of transforming DNA. In contrast to nuclear transformation ptDNA transformation of the experimental object C. reinhardtii by means of electroporation hadn´t been described. Hence a wide range of values of different physical parameters which can significantly influence the transformation efficiency was tested.
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