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New constructs and strategies for efficient PCR‐based gene manipulations in yeast
Author(s) -
Puig Oscar,
Rutz Berthold,
Luukkonen B. G. Mattias,
KandelsLewis Stefanie,
BragadoNilsson Elisabeth,
Séraphin Bertrand
Publication year - 1998
Publication title -
yeast
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.923
H-Index - 102
eISSN - 1097-0061
pISSN - 0749-503X
DOI - 10.1002/(sici)1097-0061(19980915)14:12<1139::aid-yea306>3.0.co;2-b
Subject(s) - biology , homologous recombination , plasmid , yeast , gene , genome , genetics , computational biology , dna , homologous chromosome
Gene disruption and tagging can be achieved by homologous recombination in the yeast genome. Several PCR‐based methods have been described towards this end. However these strategies are often limited in their applications and/or their efficiencies and may be technically demanding. Here we describe two plasmids for C‐terminal tagging of proteins with the IgG binding domain of the Staphyloccocus aureus protein A. We also present simple and reliable strategies based on PCR to promote efficient integration of exogenous DNA into the yeast genome. These simple methods are not limited to specific strains or markers and can be used for any application requiring homologous recombination such as gene disruption and epitope tagging. These strategies can be used for consecutive introduction of various constructs into a single yeast strain. © 1998 John Wiley & Sons, Ltd.