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  • Title: Reporter assay systems for [URE3] detection and analysis.
    Author: Brachmann A, Toombs JA, Ross ED.
    Journal: Methods; 2006 May; 39(1):35-42. PubMed ID: 16762564.
    Abstract:
    The Saccharomyces cerevisiae prion [URE3] is the infectious amyloid form of the Ure2p protein. [URE3] provides a useful model system for studying amyloid formation and stability in vivo. When grown in the presence of a good nitrogen source, [URE3] cells are able to take up ureidosuccinate, an intermediate in uracil biosynthesis, while cells lacking the [URE3] prion can not. This ability to take up ureidosuccinate has been commonly used to assay for the presence of [URE3]. However, this assay has a number of practical limitations, affecting the range of experiments that can be performed with [URE3]. Here, we describe recently developed alternative selection methods for the presence or absence of [URE3]. They make use of the Ure2p-regulated DAL5 promoter in conjunction with ADE2, URA3, kanMX, and CAN1 reporter genes, and allow for higher stringency in selection both for and against [URE3], nonselective assay of prion variants, and direct transformation of prion filaments. We discuss advantages and limitations of each of these assays.
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