for rapid creation of high-quality YKO haploids from heterozygous diploids
Genome-wide collections of yeast knockout (YKO) mutants have
facilitated the systematic functional analysis of yeast genes. An efficient
method for analyzing knockout phenotypes in parallel is to pool knockout
strains tagged with molecular barcodes and to measure their growth success in a
test culture relative to a control culture by hybridization with barcode
microarrays. However, haploid and homozygous diploid YKOs experience strong
selection for compensatory mutations and so the genetic quality of such strains
deteriorates over time. Knockout phenotypes can be protected from selection by
“hiding” the mutation as a heterozygote with the wild-type allele. However, it
is then correspondingly difficult to detect the knockout phenotype.
High-quality knockout haploids can be created from heterozygotes by tetrad
analysis, but this is labor intensive and unsuitable for genome-wide
experiments. The Magic Marker
technology (using the SGA reporter developed by Boone and coworkers1) allows fresh creation of
haploid knockouts from heterozygous diploids in a high-throughput manner by
employing a simple selection step following sporulation2. Thus, a starting pool containing thousands of high-quality MATa
haploid YKOs can be efficiently created.
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Shipping Information:
Yeast Magic Marker Strains are available as glycerol stocks.
Individual strains are shipped in 2.0‑ml tubes at room temperature; the entire
collection is shipped in 96-well plates on dry ice. All strains should be stored at –80°C.
References:
1Tong et al (2001) Science 294:2364
2Pan et al (2004) Molecular Cell 16:
487–496
