New modules for the repeated internal and N-terminal epitope tagging of genes inSaccharomyces cerevisiae

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Abstract
Epitope tagging is a powerful method for the rapid analysis of protein function. In Saccharomyces cerevisiae epitope tags are introduced easily into chromosomal loci by homologous recombination using a simple PCR‐based strategy. Although quite a number of tools exist for C‐terminal tagging as well as N‐terminal tagging of proteins expressed by heterologous promoters, there are only very limited possibilities to tag proteins at the N‐terminus and retain the endogenous expression level. Furthermore, no PCR‐templates for internal tagging have been reported. Here we describe new modules that are suitable for both the repeated N‐terminal and internal tagging of proteins, leaving their endogenous promoters intact. The tags include 6xHA, 9xMyc, yEGFP, TEV‐GST‐6xHIS, ProtA, TEV‐ProtA and TEV‐ProtA‐7xHIS in conjunction with different heterologous selection markers. Copyright © 2004 John Wiley & Sons, Ltd.