Andrie Rachael M, Martinez J Patrick, Ciuffetti Lynda M
Department of Botany and Plant Pathology, Oregon State University, Corvallis 97331, USA.
Mycologia. 2005 Sep-Oct;97(5):1152-61. doi: 10.3852/mycologia.97.5.1152.
The green fluorescent protein (GFP) has been established as the premier in vivo reporter for investigations of gene expression, protein localization, and cell and organism dynamics. The fungal transformation vector pCT74, with sGFP under the control of the ToxA promoter from Pyrenophora tritici-repentis, effectively expresses GFP in a diverse group of filamentous ascomycetes. Due to the versatility of ToxA promoter-driven expression of GFP, we constructed an additional set of fluorescent protein expression vectors to expand the color palette of fluorescent markers for use in filamentous fungi. EYFP, ECFP and mRFP1 were successfully expressed from the ToxA promoter in its fungus of origin, P. tritici-repentis, and a distant relative, Verticillium dahliae. Additionally the ToxB promoter from P. tritici-repentis drove expression of sGFP in V. dahliae, suggesting a similar potential to the ToxA promoter for heterologous expression in ascomycetes. The suite of fungal transformation vectors presented here promise to be useful for a variety of fungal research applications.
绿色荧光蛋白（GFP）已成为用于基因表达、蛋白质定位以及细胞和生物体动态研究的首要体内报告基因。真菌转化载体pCT74带有受小麦黄斑叶枯病菌（Pyrenophora tritici-repentis）ToxA启动子控制的sGFP，能在多种丝状子囊菌中有效表达GFP。由于ToxA启动子驱动的GFP表达具有通用性，我们构建了另一组荧光蛋白表达载体，以扩展用于丝状真菌的荧光标记的颜色范围。增强型黄色荧光蛋白（EYFP）、增强型青色荧光蛋白（ECFP）和单体红色荧光蛋白1（mRFP1）在其来源真菌小麦黄斑叶枯病菌以及远亲大丽轮枝菌（Verticillium dahliae）中均成功由ToxA启动子表达。此外，小麦黄斑叶枯病菌的ToxB启动子在大丽轮枝菌中驱动sGFP的表达，这表明ToxB启动子在子囊菌中进行异源表达具有与ToxA启动子类似的潜力。本文介绍的这套真菌转化载体有望用于各种真菌研究应用。
