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URL: http://www.addgene.org/44552

Proper Citation: RRID:Addgene_44552

Insert Name: PTETREG promoter

Organism: Synthetic

Bacterial Resistance: Ampicillin

Defining Citation: PMID:22511863

Vector Backbone Description: Backbone Marker:Stratagene; Backbone Size:4456; Vector Backbone:pRS403; Vector Types:Yeast Expression, Synthetic Biology, Expression regulator/reporter; Bacterial Resistance:Ampicillin

Comments: This plasmids was created as follows. First, the PTETREG promoter consisting of two tetO2 sites upstream of the minimal PCYC1 promoter was amplified from the pBB247 plasmid (Becskei, Séraphin, and Serrano, EMBO 2001, PMID: 11350942), and inserted into the pDN-G1GZmh plasmid between the AflII and BamHI sites instead of the PGAL1-D12 promoter resulting in the pDN-T2dGZmh plasmid. In order to facilitate the planned integration of the reporter plasmid into the his3Δ200 locus of the YPH500 strain, a small region bearing homology to the his3Δ200 locus was constructed by PCR and inserted in front of the HIS3 gene between the AhdI and AfeI sites of the pDN-T2dGZmh plasmid, resulting in the pDN-T2dGZmlh plasmid. Next, a second ADH1 terminator was removed from the pDN-T2dGZmlh plasmid, resulting in the final reporter pDN-T2dGZmxh plasmid bearing the yEGFP::zeoR fluorescent reporter gene under the control of the PTETREG rtTA-MF inducible promoter.

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