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.
Original language | English |
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Pages (from-to) | Unit 5.14 |
Journal | Current Protocols in Human Genetics |
Volume | Chapter 5 |
State | Published - May 2001 |
Externally published | Yes |