Cancer Research UK London Research Institute, Clare Hall Laboratories, South Mimms, UK.
Yeast. 2013 Sep;30(9):341-51. doi: 10.1002/yea.2967. Epub 2013 Jul 23.
Fusion of inducible degradation signals, so-called degrons, to cellular proteins is an elegant method of controlling protein levels in vivo. Recently, a degron system relying on the plant hormone auxin has been described for use in yeast and vertebrate cells. We now report the construction of a series of vectors that significantly enhance the versatility of this auxin-inducible degron (AID) system in Saccharomyces cerevisiae. We have minimized the size of the degron and appended a series of additional epitope tags, allowing detection by commercial antibodies or fluorescence microscopy. The vectors are compatible with PCR-based genomic tagging strategies, allow for C- or N-terminal fusion of the degron, and provide a range of selection markers. Application to a series of yeast proteins, including essential replication factors, provides evidence for a general usefulness of the system.
诱导降解信号(所谓的降解标签)与细胞蛋白融合是一种在体内控制蛋白质水平的优雅方法。最近,一种依赖植物激素生长素的降解标签系统已被描述用于酵母和脊椎动物细胞。我们现在报告了一系列载体的构建,这些载体显著增强了生长素诱导的降解标签(AID)系统在酿酒酵母中的多功能性。我们已经最小化了降解标签的大小,并添加了一系列额外的表位标签,允许通过商业抗体或荧光显微镜进行检测。这些载体与基于 PCR 的基因组标记策略兼容,允许在 C 端或 N 端融合降解标签,并提供一系列选择标记。该系统在一系列酵母蛋白(包括必需的复制因子)中的应用为其普遍适用性提供了证据。