Figure 4

Rapid epitope tagging strategy in yeast. The top line represents the target ORF in the context of a large yeast plasmid or YAC. Line 2 shows a PCR product containing the epitope tagging cassette with 40 bp targeting sequences homologous to the regions designated by the dashed lines. Line 3 shows the site-specific integration of the cassette resulting from homologous recombination in yeast. The final line represents UL114 in the YAC with an in frame 35 aa amino terminal insertion containing the ICP4 epitope and a single loxP site. This strategy can be used to place the epitope tag anywhere in the ORFs on the YAC by changing the targeting sequences on the PCR primers.