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酵母双杂交系统中蛋白质-蛋白质相互作用的联合转化与遗传技术验证

Combined transformation and genetic technique verification of protein-protein interactions in the yeast two-hybrid system.

作者信息

Tyagi S, Lal S K

机构信息

Virology Group, International Centre for Genetic Engineering and Biotechnology, New Delhi, 1100069, India.

出版信息

Biochem Biophys Res Commun. 2000 Nov 2;277(3):589-93. doi: 10.1006/bbrc.2000.3720.

DOI:10.1006/bbrc.2000.3720
PMID:11061998
Abstract

The yeast two-hybrid system is frequently used to identify protein-protein interactions. The assay is based on the functional reconstitution of a transcriptional activator. Since an indirect phenotype of the positive clones is the basis for selection of positive interacting clones, the two-hybrid screens are vulnerable to false positives. Here we report a screening protocol based on the sequential use of the cotransformation approach followed by the genetic method for verifying true two-hybrid interactions. Using this procedure, we have screened a cDNA library and have been able to isolate true positives from the yeast two-hybrid screen.

摘要

酵母双杂交系统常用于鉴定蛋白质-蛋白质相互作用。该检测基于转录激活因子的功能重建。由于阳性克隆的间接表型是选择阳性相互作用克隆的基础，双杂交筛选容易出现假阳性。在此，我们报告一种筛选方案，该方案基于共转化方法的顺序使用，随后采用遗传方法验证真正的双杂交相互作用。使用此程序，我们筛选了一个cDNA文库，并能够从酵母双杂交筛选中分离出真正的阳性克隆。

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