Shoemaker D D, Lashkari D A, Morris D, Mittmann M, Davis R W
Department of Biochemistry, Beckman Center, Stanford University Medical Center, CA 94305, USA.
Nat Genet. 1996 Dec;14(4):450-6. doi: 10.1038/ng1296-450.
A quantitative and highly parallel method for analysing deletion mutants has been developed to aid in determining the biological function of thousands of newly identified open reading frames (ORFs) in Saccharomyces cerevisiae. This approach uses a PCR targeting strategy to generate large numbers of deletion strains. Each deletion strain is labelled with a unique 20-base tag sequence that can be detected by hybridization to a high-density oligonucleotide array. The tags serve as unique identifiers (molecular bar codes) that allow analysis of large numbers of deletion strains simultaneously through selective growth conditions. Hybridization experiments show that the arrays are specific, sensitive and quantitative. A pilot study with 11 known yeast genes suggests that the method can be extended to include all of the ORFs in the yeast genome, allowing whole genome analysis with a single selective growth condition and a single hybridization.
已开发出一种用于分析缺失突变体的定量且高度并行的方法，以帮助确定酿酒酵母中数千个新鉴定的开放阅读框（ORF）的生物学功能。该方法采用PCR靶向策略来生成大量缺失菌株。每个缺失菌株都用一个独特的20碱基标签序列进行标记，该序列可通过与高密度寡核苷酸阵列杂交来检测。这些标签作为独特的标识符（分子条形码），允许通过选择性生长条件同时分析大量缺失菌株。杂交实验表明，这些阵列具有特异性、敏感性和定量性。对11个已知酵母基因的初步研究表明，该方法可扩展到涵盖酵母基因组中的所有ORF，从而在单一选择性生长条件和一次杂交的情况下实现全基因组分析。
