17_Deletion Sets

17_Deletion Sets - Strains grown for 0(red or 60...

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Unformatted text preview: Strains grown for 0 (red) or 60 generations (green) in minimal media Given a set of deletions for all genes in the genome, each can be tested and phenotyped. This is being done for budding yeast using a large number of criteria. Since at present many genes do not display a phenotype when disrupted and you can always add new selection criteria, it is an on going analysis. For whole set, 18.7% of genes are essential on rich yeast media. The information is available for every gene as part of the SGD information pages Fig. 2 from Winzeler et al (1999) When disrupting or deleting a gene by homologous recombination a unique molecular tag (DNA sequence) can be added to each locus. Homologous regions locus kanamycinR plus barcode tags Example of real tag structure (Giaever et al., 2002) Elizabeth A.Winzeler, Daniel. D. Shoemaker, Anna Astromoff, Hong Liang, et al. Functional Characterization of the Saccharomyces cerevisiae Genome by Gene Deletion and Parallel Analysis. Science. 285 : 901-906. (1999) Since each gene is uniquely tagged they can be mixed together, put under some type of genetic selection and then analyzed by microarray. Relative growth rates in rich vs minimal media for genes On chromosome XIII Fig. 3 from Winzeler et al. 1999 The complete deletion set has been tested by this means. Whole genome parallel analysis (Giaever et al., 2002 Nature. 418:387-91) Used six conditions: •High salt •Sorbitol Using highly sensitive comparison of 500 strains, 40% show quantitative growth defects. •Galactose •pH 8 •Minimal medium •Nystatin 1 Growth rates under selection Previous studies done on haploid strains. If one asks the same sorts of questions regarding heterozygous diploids then about 3% of genes generate a fitness defect. (Giaever et al., 2005) Fig.2 Giaever et al, 2002 If you couple these results to microarray expression data from stressed populations.. Plots of levels under stress vs. fitness defect under the same conditions. Fig. 4 from Giaever et al., 2002 “We found that in galactose, less than 7% of the genes that exhibited a significant increase in mRNA expression also exhibited a significant decrease in fitness. In the case of pH 8, 1 M NaCl and 1.5 M sorbitol, 3.0%, 0.88% and 0.34%, respectively, of the genes that exhibited a significant increase in mRNA expression also exhibited a significant decrease in fitness.” Giaever et al., 2002 2 ...
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