Primers Design For Yeast Integration
From 2007.igem.org
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| - | To create our | + | 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 Fv Hind CGCGGATCCATCAGTATTACCCTCGACCTC | ||
Latest revision as of 11:39, 23 October 2007
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
