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Vyučující
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Vácha František, prof. RNDr. Ph.D.
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Franta Zdeněk, RNDr. Ph.D.
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Dyčka Filip, Mgr. Ph.D.
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Fessl Tomáš, Mgr. Ph.D.
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Obsah předmětu
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1. DNA structure, history, sequencing, plasmids, cloning 2. Recombinant DNA technology, cosmids, BACs, DNA libraries 3. Gene expression in prokaryotes, promoters, expression systems, Escherichia coli 4. Microbial biotechnology, recombinant enzymes, drugs, bioremediation 5. Eukaryotic expression systems, yeast, insect cells 6. Yeast biotechnology 7. Transgenic animals, side-directed mutagenesis 8. Protein folding, design, mutagenesis, industrial enzymes 9 . Transgenic plants, methodology, Bt and HT plants 10. Transgenic plants second generation, biopharmng, regulation, public issues 11. Protein engineering: directed evolution, library preparation, display systems, applications 12. Protein purification, recombinant proteins, protein solubility, chromatographic techniques, protein tagging
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Studijní aktivity a metody výuky
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Monologická (výklad, přednáška, instruktáž), Demonstrace, Laboratorní práce
- Domácí příprava na výuku
- 100 hodin za semestr
- Účast na výuce
- 39 hodin za semestr
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Výstupy z učení
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DNA (deoxyribonucleic acid) is the carrier of genetic information in each organism. Genetic engineering, the specific and directed alteration of an organism´s hereditary material, have changed modern biology and biotechnology. There are thousands applications of using of bacteria, yeasts or insect cells of the production of recombinant proteins and various chemical compounds, many of them on the industrial scale. Transgenic crops conferring e.g. herbicide and pest resistance are another successful example of genetic engineering and new crop cultivars with modified nutritional value are just entering the market. Producing of transgenic animals for food industry is still a controversial issue but transgenic salmon has been recently approved for commercial growing. Protein engineering, a specific part of genetic engineering, is the process of developing useful or valuable proteins. It is a young discipline, with much research currently taking place into the understanding of protein folding and protein recognition for protein design principles. Both rational protein design and directed evolution techniques based on random mutagenesis are employed to generate molecules with novel properties. The course focuses on the molecular and genetic tools used to analyze and modify genetic material and to modify organisms to produce desired molecules and proteins. Topics will include sequencing techniques, cloning vectors and hosts, directed mutagenesis, and the manipulation of expression (and its levels) of particular gene products. Special attention will be directed to study biological systems utilized for the large scale production of recombinant autologous or heterologous proteins, focusing on advantages and disadvantages of each system, to allows students to evaluate and solve problems related to the expression of recombinant proteins. Furthermore, during the course, the major applications of genetic engineering in health care, forensics and agriculture will be presented. Historical overview will help students to understand present day technologies. Lectures on protein engineering will focus on preparation of gene libraries, selection of new proteins by display systems (phage, cell and cell-free) and applications (enzymes, antibodies). Students will also learn how to express and purify proteins using available techniques.
Students master theoretical knowledge about construction of genetically modified bacteria, plants and animal and about development of new proteins using protein engineering. Students have insight into Commercial applications of genetic and protein engineering including industral enzymes, transgenic crops or animals as well as into regulation connected to this field in various countries. In practical part, students acquire skills in cloning a gene into a bacterial vector, expressing and purifying recombinant proteins from Escheriachia coli and analyzing the purified proteins by immunodetection using specific antibodies.
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Předpoklady
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Basic knowledge of biochemistry, genetics and cell biology. A minimal laboratory practice - pippeting, preparation of biological buffers, calculation of solution concentrations.
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Hodnoticí metody a kritéria
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Analýza výkonů studenta
Before examination: completion of all laboratory exercises, submission of a laboratory protocol . Examination: Understanding the principles of theoretical as well as practical aspects covered in lectures and laboratory exercises. Writting exam, 50% points as a minimum
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Doporučená literatura
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Glick, R., Pasternak, J.J., Patten, C.L. Molecular Biotechnology. 2010. ISBN 978-1-55581-498-4.
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