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利用高通量筛选系统鉴定延长寿命的基因。

Identifying genes that extend life span using a high-throughput screening system.

作者信息

Chen Cuiying, Contreras Roland

机构信息

Department of Molecular Biomedical Research, Ghent University and Flanders Interuniversity Institute for Biotechnology (VIB), Ghent, Belgium.

出版信息

Methods Mol Biol. 2007;371:237-48. doi: 10.1007/978-1-59745-361-5_18.

Abstract

We developed a high-throughput functional genomic screening system that allows identification of genes prolonging lifespan in the baker's yeast Saccharomyces cerevisiae. The method is based on isolating yeast mother cells with a higher than average number of cell divisions as indicated by the number of bud scars on their surface. Fluorescently labeled wheat germ agglutinin (WGA) was used for specific staining of chitin, a major component of bud scars. The critical new steps in our bud-scar-sorting system are the use of small microbeads, which allows successive rounds of purification and regrowth of the mother cells (M-cell), and utilization of flow cytometry to sort and isolate cells with a longer lifespan based on the number of bud scars specifically labeled with WGA.

摘要

我们开发了一种高通量功能基因组筛选系统,该系统能够鉴定出延长酿酒酵母寿命的基因。该方法基于分离酵母母细胞,这些母细胞表面芽痕数量表明其细胞分裂次数高于平均水平。荧光标记的小麦胚凝集素(WGA)用于对芽痕的主要成分几丁质进行特异性染色。我们的芽痕分选系统中的关键新步骤是使用小微珠,这使得母细胞(M细胞)能够进行连续多轮的纯化和再生长,以及利用流式细胞术根据被WGA特异性标记的芽痕数量来分选和分离寿命更长的细胞。

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