Willer M, Regnacq M, Reid P J, Tyson J R, Cui W, Wilkinson B M, Stirling C J
School of Biological Sciences, 2.205 Stopford Building, University of Manchester, Oxford Road, Manchester M13 9PT, UK.
Yeast. 2000 Nov;16(15):1429-35. doi: 10.1002/1097-0061(200011)16:15<1429::AID-YEA629>3.0.CO;2-S.
In the context of the EUROFAN programme, we report the deletion and functional analysis of six open reading frames (ORFs) on the right arm of chromosome XII of Saccharomyces cerevisiae. Using a PCR-based gene replacement strategy, we have systematically deleted individual ORFs and subjected the heterozygous diploids and haploid knockout strains to basic genetic and phenotypic characterization. Two ORFs, YLR127c and YLR129w, are essential for viability, whereas no growth phenotype could be detected following deletion of YLR124w, YLR125w, YLR126c or YLR128w. For each of the individual ORFs, a kanMX4 replacement cassette and the corresponding cognate wild-type gene were cloned into appropriate plasmids.
在EUROFAN项目的背景下，我们报告了酿酒酵母第十二号染色体右臂上六个开放阅读框（ORF）的缺失及功能分析。利用基于聚合酶链反应（PCR）的基因替换策略，我们系统地删除了各个ORF，并对杂合二倍体和单倍体基因敲除菌株进行了基本的遗传学和表型特征分析。两个ORF，YLR127c和YLR129w，对细胞活力至关重要，而删除YLR124w、YLR125w、YLR126c或YLR128w后未检测到生长表型。对于每个单独的ORF，将一个kanMX4替换盒和相应的同源野生型基因克隆到合适的质粒中。
