Tagging strains using Longtine plasmids

Introduction

This method allows quick deletion or tagging of any gene in yeast using PCR. This method is a quick summary of the original paper by Mark S. Longtine et al., Yeast 14, 953-961 (1998).

Design Primers


F1 5'-(gene-specific sequence)-CGGATCCCCGGGTTAATTAA Deletion
R1 5'-(gene-specific sequence)-GAATTCGAGCTCGTTTAAAC Deletion/C-terminal Tagging
F2 5'-(gene-specific sequence)-CGGATCCCCGGGTTAATTAA C-terminal tagging

Also design two primers to check for correct insertion. One internal, one external to the gene of interest.