From 2007.igem.org

To create our circuit in yeast cell, we have to integrate '2XORE' promoter in place of the Pho4 endogenous promoter.

We cloned 2XORE into Hind-Bam E.R. site of p2UG25 plasmid that has natMX4 resistence, thus contructing a 2XORE-natMX4 cassette,which we can integrate unpstream Pho4 ORF.

2XORE Fv Hind CGCGGATCCATCAGTATTACCCTCGACCTC

2XORE Rv Bam CCCAAGCTTTGTTTGCGTGTCTATAGAAG

2XOREint Fv TCTAGATCGTCCACAAAACCGTGTATTCCCTCAGAAGTTGTACGGCCCATTGTTTGCGTGTCTATAGAAG

2XOREint Rv CAGAAGAGAAGAGATGAGCAAAGGAGACAGAACAAGAGTAGCAGAAAGTCATCGATGAATTCGAGCTCG

We tagged Pho8gene with GFP,integrating GFP-KanMX6 cassette in Pho8's ORF

Fv TTACATATCAGCATACGGGACATTATTTGAACGCGCATTAGCAGCATGAGTAAAGGAGAAGAACTTTTC

Rv TCAGATCCAGGAACAAGACGTGTCTGTTCGCTTGGTAATGTGTGAGTCATATCGATGAATTCGAGCTCG

To delete Pho80 endogenous and Pho81, we integrate into their CDS hphMX4 and Kltrip resistence,respective.

hphMX4-Pho80Fv ATGGAAAGCACATCAGGAGAACGTTCCGAAAATATACATGAGGATCAAGGCGTACGCTGCAGGTCGAC

hphMX4-Pho80Rv TTAATCTGGCTTTGATCGCTTGTTATAGATATGTGCGTCCTTTGCCTCAGATCGATGAATTCGAGCTCG

Kltrip-Pho81Fv ATGAAATTCGGCAAGTATTTGGAAGCCAGGCAGTTGGAACTGGCGGAGTAACTAGCTAAAAAGTGGAACGA

Kltrip-Pho81Rv TTACATATCAATATTATTCTCAAAAAGCAACTCACAAGCGTAGTAGATCCCTCGAGGCCAGAAGACT