Zhou Xiaoying, Li Guotian, Xu Jin-Rong
Department of Botany and Plant Pathology, Purdue University, West Lafayette, IN, USA.
Methods Mol Biol. 2011;722:199-212. doi: 10.1007/978-1-61779-040-9_15.
For functional characterization of predicted genes encoding hypothetical proteins in fungal genomes, it is complementary to genetic studies to determine their expression and subcellular localization patterns in different developmental or infection stages. It is also important to identify and characterize other proteins that are physically associated with or functionally related to these genes in vivo by co-immunoprecipitation or affinity purification analyses. In this chapter, we described a set of yeast shuttle vectors and protocols to generate fusion constructs by the yeast gap repair approach. Because of the simplicity and efficiency of yeast gap repair, these vectors and the general methods described in this chapter are suitable for functional genomics studies in filamentous fungi.
对于真菌基因组中编码假定蛋白质的预测基因进行功能表征，确定其在不同发育或感染阶段的表达和亚细胞定位模式，是对遗传学研究的补充。通过免疫共沉淀或亲和纯化分析，在体内鉴定和表征与这些基因存在物理关联或功能相关的其他蛋白质也很重要。在本章中，我们描述了一组酵母穿梭载体和通过酵母缺口修复方法生成融合构建体的方案。由于酵母缺口修复的简便性和高效性，本章所述的这些载体和通用方法适用于丝状真菌的功能基因组学研究。
