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Curr Protoc Hum Genet. 2001 May;Chapter 5:Unit 5.14. doi: 10.1002/0471142905.hg0514s06.

Transfer of YAC clones to new hosts by karyogamy-deficient mating.

Author information

1
Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.

Abstract

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.

PMID:
18428288
DOI:
10.1002/0471142905.hg0514s06
[Indexed for MEDLINE]

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