Transfer of YAC clones to new hosts by karyogamy-deficient mating.
This unit provides a protocol for moving yeast artificial chromosome (YAC) clones to new yeast hosts using basic microbial techniques and pulsed-field gel analysis. In contrast to other methods that can be used to transfer YAC clones, this requires neither optimization to achieve high-efficiency DNA-mediated transformation of chromosome-sized DNA nor specialized equipment for tetrad dissection and analysis. Instead, chromosome (YAC) transfer is selected in rare segregants ("YACductants") from a yeast mating that is rendered incomplete in most cell pairings by the presence of a kar1 (karyogamy-deficient) mutation in either parental strain. The Basic Protocol in this unit details the transfer of a YAC clone from yeast strain AB1380 (host to nearly all existing YAC libraries) to YPH925, a strain with nonreverting genetic markers compatible with existing plasmid constructs useful in YAC modification.
Spencer
F
F
Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
Simchen
G
G
D018244:Chromosomes, Artificial, Yeast / Q000235:genetics*; D005800:Genes, Fungal; D005821:Genetic Techniques; D005826:Genetics, Medical; D006801:Humans; D007621:Karyotyping; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics
C074559:KAR1 protein, S cerevisiae; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins