. . . . "M\u00FCller, Petr" . "Metody exprese a purifikace rekombinantn\u00EDch protein\u016F umo\u017E\u0148uj\u00ED produkci a detailn\u00ED charakterizaci protein\u016F v z\u00E1kladn\u00EDm v\u00FDzkumu b\u011Bhem in vitro experiment\u016F, ale tak\u00E9 p\u0159\u00EDpravu protein\u016F s terapeutick\u00FDm vyu\u017Eit\u00EDm. Publikace shrnuje z\u00E1kladn\u00ED postupy od p\u0159\u00EDpravy expresn\u00EDch vektor\u016F a\u017E po techniku afinitn\u00ED purifikace. D\u00E1le pojedn\u00E1v\u00E1 o vlastnostech r\u016Fzn\u00FDch prokaryotick\u00FDch a eukaryotick\u00FDch expresn\u00EDch syst\u00E9m\u016F a mo\u017Enostech jejich vyu\u017Eit\u00ED. Molekul\u00E1rn\u00ED klonov\u00E1n\u00ED, kter\u00E9 slou\u017E\u00ED k p\u0159\u00EDprav\u011B expresn\u00EDch vektor\u016F pro rekombinantn\u00ED proteiny, umo\u017E\u0148uje c\u00EDlen\u011B modifikovat vlastnosti t\u011Bchto protein\u016F tak, aby byla usnadn\u011Bna jejich purifi kace a tak\u00E9 pozm\u011Bn\u011Bna jejich stabilita, aktivita nebo funkce. V sou\u010Dasn\u00E9 dob\u011B je k dispozici \u0161irok\u00E1 \u0161k\u00E1la metodick\u00FDch p\u0159\u00EDstup\u016F, je\u017E umo\u017E\u0148uj\u00ED rychlou a efektivn\u00ED p\u0159\u00EDpravu expresn\u00EDch vektor\u016F. Zvolen\u00FD produk\u010Dn\u00ED organizmus a zp\u016Fsob purifikace rekombinantn\u00EDho proteinu ur\u010Duj\u00ED v\u00FDb\u011Br expresn\u00EDho vektoru. Prvn\u00ED volbou \u010Dasto b\u00FDv\u00E1 expresn\u00ED syst\u00E9m vyu\u017E\u00EDvaj\u00EDc\u00ED bakterii Escherichia coli, jeho\u017E p\u0159ednostmi jsou zejm\u00E9na technick\u00E1, \u010Dasov\u00E1 i finan\u010Dn\u00ED nen\u00E1ro\u010Dnost. Tento expresn\u00ED syst\u00E9m nen\u00ED p\u0159\u00EDli\u0161 vhodn\u00FD pro produkci komplexn\u00EDch sav\u010D\u00EDch protein\u016F, pro kter\u00E9 jsou optim\u00E1ln\u00ED expresn\u00ED syst\u00E9my zalo\u017Een\u00E9 na vyu\u017Eit\u00ED eukaryotick\u00FDch organizm\u016F (kvasinky, hmyz\u00ED bu\u0148ky nebo sav\u010D\u00ED bu\u0148ky). Kultivace hmyz\u00EDch a sav\u010D\u00EDch bun\u011Bk je v\u0161ak technicky i finan\u010Dn\u011B n\u00E1ro\u010Dn\u00E1. Rekombinantn\u00ED proteiny jsou purifikov\u00E1ny nej\u010Dast\u011Bji metodou afinitn\u00ED chromatografie vyu\u017E\u00EDvaj\u00EDc\u00ED specifickou interakci peptidu nebo proteinu s afinitn\u00ED matric\u00ED. Tyto peptidy \u010Di proteiny jsou f\u00FAzov\u00E1ny s N- nebo C-koncem purifikovan\u00E9ho proteinu. Purifikace prob\u00EDh\u00E1 ve t\u0159ech kroc\u00EDch, kdy je rekombinantn\u00ED protein prost\u0159ednictv\u00EDm afinitn\u00EDch zna\u010Dek specificky zachycen na matrici chromatografick\u00E9 kolony, d\u00E1le n\u00E1sleduje prom\u00FDvac\u00ED krok, po kter\u00E9m je uvoln\u011Bn z kolony \u010Dist\u00FD protein."@cs . "RIV/00209805:_____/14:#0000535" . "16136" . "3"^^ . "Exprese a purifikace protein\u016F" . . "CZ - \u010Cesk\u00E1 republika" . "0862-495X" . "Exprese a purifikace protein\u016F"@cs . . "Vojt\u011B\u0161ek, Bo\u0159ivoj" . . "recombinant protein; molecular cloning; purification; expression system"@en . . "I, P(ED2.1.00/03.0101)" . . "Protein expression and purification"@en . . "27" . "6"^^ . . . "RIV/00209805:_____/14:#0000535!RIV15-MSM-00209805" . . "Klinick\u00E1 onkologie" . "Exprese a purifikace protein\u016F" . "supplementum 1" . . . . "Exprese a purifikace protein\u016F"@cs . "[D58BFB37DF1E]" . "Production of recombinant proteins is essential for many applications in both basic research and also in medicine, where recombinant proteins are used as pharmaceuticals. This review summarizes procedures involved in recombinant protein expression and purification, including molecular cloning of target genes into expression vectors, selection of the appropriate expression system, and protein purification techniques. Recombinant DNA technology allows protein engineering to modify protein stability, activity and function or to facilitate protein purification by affinity tag fusions. A wide range of cloning systems enabling fast and effective design of expression vectors is currently available. A first choice of protein expression system is usually the bacteria Escherichia coli. The main advantages of this prokaryotic expression system are low cost and simplicity; on the other hand this system is often unsuitable for production of complex mammalian proteins. Protein expression mediated by eukaryotic cells (yeast, insect and mammalian cells) usually produces properly folded and posttranslationally modified proteins. However, cultivation of insect and, especially, mammalian cells is time consuming and expensive. Affinity tagged recombinant proteins are purified efficiently using affinity chromatography. An affinity tag is a protein or peptide that mediates specific binding to a chromatography column, unbound proteins are removed during a washing step and pure protein is subsequently eluted."@en . . . "R\u016F\u010Dkov\u00E1, Eva" . . "http://www.linkos.cz/files/klinicka-onkologie/186/4493.pdf" . "Protein expression and purification"@en . "Metody exprese a purifikace rekombinantn\u00EDch protein\u016F umo\u017E\u0148uj\u00ED produkci a detailn\u00ED charakterizaci protein\u016F v z\u00E1kladn\u00EDm v\u00FDzkumu b\u011Bhem in vitro experiment\u016F, ale tak\u00E9 p\u0159\u00EDpravu protein\u016F s terapeutick\u00FDm vyu\u017Eit\u00EDm. Publikace shrnuje z\u00E1kladn\u00ED postupy od p\u0159\u00EDpravy expresn\u00EDch vektor\u016F a\u017E po techniku afinitn\u00ED purifikace. D\u00E1le pojedn\u00E1v\u00E1 o vlastnostech r\u016Fzn\u00FDch prokaryotick\u00FDch a eukaryotick\u00FDch expresn\u00EDch syst\u00E9m\u016F a mo\u017Enostech jejich vyu\u017Eit\u00ED. Molekul\u00E1rn\u00ED klonov\u00E1n\u00ED, kter\u00E9 slou\u017E\u00ED k p\u0159\u00EDprav\u011B expresn\u00EDch vektor\u016F pro rekombinantn\u00ED proteiny, umo\u017E\u0148uje c\u00EDlen\u011B modifikovat vlastnosti t\u011Bchto protein\u016F tak, aby byla usnadn\u011Bna jejich purifi kace a tak\u00E9 pozm\u011Bn\u011Bna jejich stabilita, aktivita nebo funkce. V sou\u010Dasn\u00E9 dob\u011B je k dispozici \u0161irok\u00E1 \u0161k\u00E1la metodick\u00FDch p\u0159\u00EDstup\u016F, je\u017E umo\u017E\u0148uj\u00ED rychlou a efektivn\u00ED p\u0159\u00EDpravu expresn\u00EDch vektor\u016F. Zvolen\u00FD produk\u010Dn\u00ED organizmus a zp\u016Fsob purifikace rekombinantn\u00EDho proteinu ur\u010Duj\u00ED v\u00FDb\u011Br expresn\u00EDho vektoru. Prvn\u00ED volbou \u010Dasto b\u00FDv\u00E1 expresn\u00ED syst\u00E9m vyu\u017E\u00EDvaj\u00EDc\u00ED bakterii Escherichia coli, jeho\u017E p\u0159ednostmi jsou zejm\u00E9na technick\u00E1, \u010Dasov\u00E1 i finan\u010Dn\u00ED nen\u00E1ro\u010Dnost. Tento expresn\u00ED syst\u00E9m nen\u00ED p\u0159\u00EDli\u0161 vhodn\u00FD pro produkci komplexn\u00EDch sav\u010D\u00EDch protein\u016F, pro kter\u00E9 jsou optim\u00E1ln\u00ED expresn\u00ED syst\u00E9my zalo\u017Een\u00E9 na vyu\u017Eit\u00ED eukaryotick\u00FDch organizm\u016F (kvasinky, hmyz\u00ED bu\u0148ky nebo sav\u010D\u00ED bu\u0148ky). Kultivace hmyz\u00EDch a sav\u010D\u00EDch bun\u011Bk je v\u0161ak technicky i finan\u010Dn\u011B n\u00E1ro\u010Dn\u00E1. Rekombinantn\u00ED proteiny jsou purifikov\u00E1ny nej\u010Dast\u011Bji metodou afinitn\u00ED chromatografie vyu\u017E\u00EDvaj\u00EDc\u00ED specifickou interakci peptidu nebo proteinu s afinitn\u00ED matric\u00ED. Tyto peptidy \u010Di proteiny jsou f\u00FAzov\u00E1ny s N- nebo C-koncem purifikovan\u00E9ho proteinu. Purifikace prob\u00EDh\u00E1 ve t\u0159ech kroc\u00EDch, kdy je rekombinantn\u00ED protein prost\u0159ednictv\u00EDm afinitn\u00EDch zna\u010Dek specificky zachycen na matrici chromatografick\u00E9 kolony, d\u00E1le n\u00E1sleduje prom\u00FDvac\u00ED krok, po kter\u00E9m je uvoln\u011Bn z kolony \u010Dist\u00FD protein." . "3"^^ .