Geymonat Marco, Spanos Adonis, Sedgwick Steven
Stem Cell Biology and Developmental Genetics, National Institute for Medical Research, London, UK.
Methods Mol Biol. 2009;545:63-80. doi: 10.1007/978-1-60327-993-2_4.
A novel protein expression system in budding yeast is described which has been used to express many yeast mitotic regulators as well as a wide range of other recombinant proteins from several different species. The expression system relies on autoselection with essential genes to maintain high copy numbers of expression plasmids. Autoselection permits expression cells to be grown in rich medium with no need for plasmid selection with drugs or nutritional conditions. This optimizes growth and expression of recombinant proteins. The use of the expression system is illustrated by purifying budding yeast mitotic regulators, Cdc14 and Net1, and recapitulating their activities in vitro.
本文描述了一种用于芽殖酵母的新型蛋白质表达系统，该系统已被用于表达多种酵母有丝分裂调节因子以及来自几个不同物种的多种其他重组蛋白。该表达系统依靠与必需基因的自动选择来维持表达质粒的高拷贝数。自动选择使表达细胞能够在丰富培养基中生长，而无需通过药物或营养条件进行质粒选择。这优化了重组蛋白的生长和表达。通过纯化芽殖酵母有丝分裂调节因子Cdc14和Net1并在体外重现它们的活性，说明了该表达系统的用途。
