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. Author manuscript; available in PMC: 2011 Mar 1.
Published in final edited form as: Fungal Genet Biol. 2009 Nov 26;47(3):191–198. doi: 10.1016/j.fgb.2009.11.007

Figure 1.

Figure 1

G418 marker recycling strategy. A. Flanking loxP sites (triangles) were added to the 2.7 kb G418 drug marker and ligated into a plasmid to create vector pJL517. B. Vector pJL517 and genomic DNA were used as templates in an overlap PCR that resulted in loxP-G418-loxP cassette flanked by the 5′ and 3′ UTR of LAC1. The native LAC1 (4.6 kb between primers #7 and #8) was replaced through homologous recombination with this construct. C. The strain created in B. was transformed with the uncut plasmid, pJL519 (CRE-recombinase); CRE was transiently expressed by growth on galactose induction medium. Translated Cre-recombinase (ovals) binds to the two-loxP sites. The 2.7 kb G418 construct is looped out, resulting in a PCR product that is 2.0 kb.