IGEM:UCSF/2007/Plasmids
From 2007.igem.org
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<u>'''PROJECT 2</u> | <u>'''PROJECT 2</u> | ||
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| + | (1) PspOMI//pSte2-Ste2//XhoI//GFP//XmaI//RR//STOP//BamHI//AdhTerminator | ||
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| + | GGGCCCATCCAATATCACCTGACCTTCATCTTTATGTTTTTCCACTGGTGTAATGTACTCTGTTGCATCATCAATAAGCAACTGGGACGCCAATAATTCATACCAACAATCTTTTCTTAATGAGTTGTTTACAAACCCCAGAGGCCCTAAACCTATGTACCTCAAAGCTCGGACATCTTTTTTCTCTAATAACTTTGATATAACTTTTCCTTTTTGTTCTTCTGCCTTTCCACTGTCACAACGAGAAAGTGATTCCAGATATGCGTTATAACCTCCCCTTGTCAACGCGTCTTTGTCGTGGCTACTCTGATTAGTATGAAATAATGACCTTAATGGCATCGTATTTTGTTAATTGGCTTGTTCTATGTCATTTGCTAAATTTTCTCTTTATTTCTTTACTTTGCCAATTCAAGATAATCTTCCCTTCCCAGAGAGAAAAAAGGAAAAATTTAGCTATGAAACCTCAATAAGCTTTTTAATACACCAAAGATTCAAGATAAGAGCATAGAACGAACTGTAGAATAGTCCGGATATGTTATCCAATGCCTGCCAAAATGCATTGTCACACGCTGTAGTGCTCGAATAGGTGTTGCAATCCGTCAATATACGTCTTGCTCTGTGGGTAAATGTCTCGTGCATTAAGACAGGCTAGTATAAACGAGAAGAAGTATCCTGCTTTGCAATGAAACAATAGTATCCGCTAAGAATTTAAGCAGGCCAACGTCCATACTGCTTAGGACCTGTGCCTGGCAAGTCGCAGATTGAAGTTTTTTCAACCATGTAAATTTCCTAATTGGGTAAGTACATGATGAAACACATATGAAGAAAAAAGCTTTCCTACATATTCAAGATTTTTTTCTGTGGGTGGAATACTATTTAAGGAGTGCTATTAGTATCTTATTTGACTTCAAAGCAATACGATACCTTTTCTTTTCACCTGCTCTGGCTATAATTATAATTGGTTACTTAAAAATGCACCGTTAAGAACCATATCCAAGAATCAAAAATGTCTGATGCGGCTCCTTCATTGAGCAATCTATTTTATGATCCAACGTATAATCCTGGTCAAAGCACCATTAACTACACTTCCATATATGGGAATGGATCTACCATCACTTTCGATGAGTTGCAAGGTTTAGTTAACAGTACTGTTACTCAGGCCATTATGTTTGGTGTCAGATGTGGTGCAGCTGCTTTGACTTTGATTGTCATGTGGATGACATCGAGAAGCAGAAAAACGCCGATTTTCATTATCAACCAAGTTTCATTGTTTTTAATCATTTTGCATTCTGCACTCTATTTTAAATATTTACTGTCTAATTACTCTTCAGTGACTTACGCTCTCACCGGATTTCCTCAGTTCATCAGTAGAGGTGACGTTCATGTTTATGGTGCTACAAATATAATTCAAGTCCTTCTTGTGGCTTCTATTGAGACTTCACTGGTGTTTCAGATAAAAGTTATTTTCACAGGCGACAACTTCAAAAGGATAGGTTTGATGCTGACGTCGATATCTTTCACTTTAGGGATTGCTACAGTTACCATGTATTTTGTAAGCGCTGTTAAAGGTATGATTGTGACTTATAATGATGTTAGTGCCACCCAAGATAAATACTTCAATGCATCCACAATTTTACTTGCATCCTCAATAAACTTTATGTCATTTGTCCTGGTAGTTAAATTGATTTTAGCTATTAGATCAAGAAGATTCCTTGGTCTCAAGCAGTTCGATAGTTTCCATATTTTACTCATAATGTCATGTCAATCTTTGTTGGTTCCATCGATAATATTCATCCTCGCATACAGTTTGGAACCAAACCAGGGAACAGATGTCTTGACTACTGTTGCAACATTACTTGCTGTATTGTCTTTACCATTATCATCAATGTGGGCCACGGCTGCTAATAATGCATCCAAAACAAACACAATTACTTCAGACTTTACAACATCCACAGATAGGTTTTATCCAGGCACGCTGTCTAGCTTTCAAACTGATAGTATCAACAACGATGCTAAAAGCAGTCTCAGAAGTAGATTATATGACCTATATCCTAGAAGGAAGGAAACAACATCGGATAAACATTCGGAAAGAACTTTTGTTTCTGAGACTGCAGATGATATAGAGAAAAATCAGTTTTATCAGTTGCCCACACCTACGAGTTCAAAAAATACTAGGATAGGACCGTTTGCTGATGCAAGTTACAAAGAGGGAGAAGTTGAACCCGTCGACATGTACACTCCCGATACGGCAGCTGATGAGGAAGCCAGAAAGTTCTGGACTGAAGATAATAATAATTTACTCGAGTCTAAAGGTGAAGAATTATTCACTGGTGTTGTCCCAATTTTGGTTGAATTAGATGGTGATGTTAATGGTCACAAATTTTCTGTCTCCGGTGAAGGTGAAGGTGATGCTACTTACGGTAAATTGACCTTAAAATTTATTTGTACTACTGGTAAATTGCCAGTTCCATGGCCAACCTTAGTCACTACTTTAACTTATGGTGTTCAATGTTTTTCTAGATACCCAGATCATATGAAACAACATGACTTTTTCAAGTCTGCCATGCCAGAAGGTTATGTTCAAGAAAGAACTATTTTTTTCAAAGATGACGGTAACTACAAGACCAGAGCTGAAGTCAAGTTTGAAGGTGATACCTTAGTTAATAGAATCGAATTAAAAGGTATTGATTTTAAAGAAGATGGTAACATTTTAGGTCACAAATTGGAATACAACTATAACTCTCACAATGTTTACATCATGGCTGACAAACAAAAGAATGGTATCAAAGTTAACTTCAAAATTAGACACAACATTGAAGATGGTTCTGTTCAATTAGCTGACCATTATCAACAAAATACTCCAATTGGTGATGGTCCAGTCTTGTTACCAGACAACCATTACTTATCCACTCAATCTaggTTATCCAAAGATCCAAACGAAAAGAGAGACCACATGGTCTTGTTAGAATTTGTTACTGCTGCTGGTATTACCCATGGTATGGATGAATTGTACAAACCCGGGggatctggatcgggatctGATCCAGATTTGGAAATTAGAGCTGCTTTTTTGAGACAAAGAAATACTGCTTTGAGAACTGAAGTTGCTGAATTGGAACAAGAAGTTCAAAGATTGGAAAATGAAGTTTCTCAATATGAAACTAGATATGGTCCATTGGGTGGTGGTAAATAAGGATCCGCGAATTTCTTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTATACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGTAACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT ATTGACCACACC | ||
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| + | (2) PspOMI//pGal1//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator | ||
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| + | TGTTATTGTGATAGTATCTCAGCGGGATTTGTGTAGATTCACCTACAGCCTTTAAACCAGCAAAGTGAATTACCGAATCAATTTTATATTCTTTGAAAACCTTTTCCAGACCTTTTCGGTCACACAAATCAACCTCATAGAAGGGAATGTGATGCTTGGTCAAGACCTCTAACCTGGCTACAGAATCATAAGTTGAATTCGACAGGTTATCAGCAACAACACAGTCATATCCATTCTCAATTAGCTCTACCACAGTGTGTGAACCAATGTATCCAGCACCACCTGTAACCAAAACAATTTTAGAAGTACTTTCACTTTGTAACTGAGCTGTCATTTATATTGAATTTTCAAAAATTCTTACTTTTTTTTTGGATGGACGCAAAGAAGTTTAATAATCATATTACATGGCATTACCACCATATACATATCCATATCTAATCTTACTTATATGTTGTGGAAATGTAAAGAGCCCCATTATCTTAGCCTAAAAAAACCTTCTCTTTGGAACTTTCAGTAATACGCTTAACTGCTCATTGCTATATTGAAGTACGGATTAGAAGCCGCCGAGCGGGCGACAGCCCTCCGACGGAAGACTCTCCTCCGTGCGTCCTCGTCTTCACCGGTCGCGTTCCTGAAACGCAGATGTGCCTCGCGCCGCACTGCTCCGAACAATAAAGATTCTACAATACTAGCTTTTATGGTTATGAAGAGGAAAAATTGGCAGTAACCTGGCCCCACAAACCTTCAAATTAACGAATCAAATTAACAACCATAGGATGATAATGCGATTAGTTTTTTAGCCTTATTTCTGGGGTAATTAATCAGCGAAGCGATGATTTTTGATCTATTAACAGATATATAAATGGAAAAGCTGCATAACCACTTTAACTAATACTTTCAACATTTTCAGTTTGTATTACTTCTTATTCAAATGTCATAAAAGTATCAACAAAAAATTGTTAATATACCTCTATACTTTAACGTCAAGGAGAAAAAACTATAATGCTCGAGGGAGCTgcttctgcatcaacttctaaatataattcacactccttggagaatgagtctattaagaggacgtctcgagatggagtcaatcgagatctcactgaggctgttcctcgacttccaggagaaacactaatcactgacaaagaagttatttacatatgtcctttcaatggccccattaagggaagagtttacatcacaaattatcgtctttatttaagaagtttggaaacggattcttctctaatacttgatgttcctctgggtgtgatctcgagaattgaaaaaatgggaggcgcgacaagtagaggagaaaattcctatggtctagatattacttgtaaagacatgagaaacctgaggttcgctttgaaacaggaaggccacagcagaagagatatgtttgagatcctcacgagatacgcgtttcccctggctcacagtctgccattatttgcatttttaaatgaagaaaagtttaacgtggatggatggacagtttacaatccagtggaagaatacaggaggcagggcttgcccaatcaccattggagaataacttttattaataagtgctatgagctctgtgacacttaccctgctcttttggtggttccgtatcgtgcctcagatgatgacctccggagagttgcaacttttaggtcccgaaatcgaattccagtgctgtcatggattcatccagaaaataagacggtcattgtgcgttgcagtcagcctcttgtcggtatgagtgggaaacgaaataaagatgatgagaaatatctcgatgttatcagggagactaataaacaaatttctaaactcaccatttatgatgcaagacccagcgtaaatgcagtggccaacaaggcaacaggaggaggatatgaaagtgatgatgcatatcataacgccgaacttttcttcttagacattcataatattcatgttatgcgggaatctttaaaaaaagtgaaggacattgtttatcctaatgtagaagaatctcattggttgtccagtttggagtctactcattggttagaacatatcaagctcgttttgacaggagccattcaagtagcagacaaagtttcttcagggaagagttcagtgcttgtgcattgcagtgacggatgggacaggactgctcagctgacatccttggccatgctgatgttggatagcttctataggagcattgaagggttcgaaatactggtacaaaaagaatggataagttttggacataaatttgcatctcgaataggtcatggtgataaaaaccacaccgatgctgaccgttctcctatttttctccagtttattgattgtgtgtggcaaatgtcaaaacagttccctacagcttttgaattcaatgaacaatttttgattataattttggatcatctgtatagttgccgatttggtactttcttattcaactgtgaatctgctcgagaaagacagaaggttacagaaaggactgtttctttatggtcactgataaacagtaataaagaaaaattcaaaaaccccttctatactaaagaaatcaatcgagttttatatccagttgccagtatgcgtcacttggaactctgggtgaattactacattagatggaaccccaggatcaagcaacaacagccgaatccagtggagcagcgttacatggagctcttagccttacgcgacgaatacataaagcggcttgaggaactgcagctcgccaactctgccaagctttctgatcccccaacttcaccttccagtccttcgcaaatgatgccccatgtgcaaactcacttcGGTAGTTCCCTAggttccggtagcggttccggttctatcactattagggctgcttttttggaaaaagaaaatactgctttgagaactgaaattgctgaattggaaaaagaagttggtagatgtgaaaatattgtttctaaatatgaaactagatatggtccattgTAATCGCGGATCCGCGAATTTCTTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTATACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGTAACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT ATTGACCACACC | ||
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| + | (3) pGal1//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator | ||
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| + | As above, but WMTM1 sequence | ||
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| + | (4) pSte5//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator | ||
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| + | As above, but Ste5 promoter sequence: | ||
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| + | CACGAAGTGACTGACAGAATACTGACATCAGCTGATTTCTCATAGAGCTGTTTCTCTGATAACACGTTGTTTGAACATCGACAAGATGAAAATCTAGAAGTATCAAGTTTCCTTTAAAGGGATATATAACAGATTCTAAAACTGACAGAAATATTTCGAGTGAAGAAGAAGCGTTAAATATTGGATCTTTCCGCAGTTCTACTCTGATACATTTTTGAAGTAGGAGAGTCATTTAGAAGGCGTATTGCTCAATAGTAGAAAGCAGGCCTGTGCACATGAATTAATTAAAAAATATAAAGGTAGTGATTAGACGACACATGTCCATAGGTAACCTGTCATAATTTTGAACAATTTCCCTTCTTTTCTTTTTTTTTTTTGGGTGCGGCGATATGTAGCTTGTTAATTTACACATCATGTACTTTTCTGCATCAAAATATGAAAGGCGATAGTAGCTAAAGAAAATACCGAGAATTTCCTCGAAAAGTTGACGACAAAAGAAAGGCATAAAAAAGTAATTTGAAAATATTTTAAAACTGTTTTAACCCATCTAGCATCCGCGCTAAAAAAGGAAGATACAGGATACAGCGGAAACAACTTTTAAATG | ||
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| + | (5) pSte5//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator | ||
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| + | As above. | ||
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| + | (6) pCyc//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator | ||
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| + | As above, but Cyc promoter sequence: | ||
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| + | ctcgagcagatccgccaggcgtgtatatatagcgtggatggccaggcaactttagtgctgacacatacaggcatatatatatgtgtgcgacgacacatgatcatatggcatgcatgtgctctgtatgtatataaaactcttgttttcttcttttctctaaatattctttccttatacattaggacctttgcagcataaattactatacttctatagacacacaaacacaaatacacacactaaattaataatg | ||
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| + | (7) pCyc//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator | ||
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| + | As above. | ||
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| + | (8) pCyc//ATG//XhoI//GFP//XmaI//(SerGlySerGlySer)ING2 PH domain//STOP//BamHI | ||
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| + | As above, but //(SerGlySerGlySer)ING2 PH domain sequence as follows: | ||
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| + | TCTGGATCAGGTTCAGCTTCTCCAGTTGAATTTGCTATTGATCCTAACGAACCTACATACTGCTTATGTAATCAAGTTTCTTATGGTGAAATGATTGGTTGCGATAACGAGCAGTGTCCAATAGAATGGTTTCATTTCTCTTGCGTTTCTTTAACATATAAACCTAAGGGAAAATGGTACTGTCCTAAATGTAGAGGTGATAATGAGAAAACTATGGATAAGTCTACTGAAAAGACAAAAAAAGATAGAAGATCAAGA | ||
Revision as of 20:08, 26 October 2007
PROJECT 1
Expected sequences
Below you have the sequences of the different parts in the right order, as well as the adaptor sequences between parts. You will need to arrange them accordingly to your particular constructs, so that you will have 1-2-3-4-5-6-7-8.
1-Promoter sequence (variable, depends on particular acceptor)
pFig1
TTGAATGATCAACCAAACGCCGATATAGTCACTATTGGTGATAAAATTAAAATGC AGCTATTGAGAACCCATCCTAACAACATTAAATTGTCTATTAATGGAATTGGTAA AGATGCCCCAACTTTCCAAGAATCTGCTTTGATTGCCGATAAGTTATTGAGTGTC ATGAAGGCCGGCACTTACCCAAAGATTTCCATTTTCTACAATGACCCAGTGTCTT CCCTATCTTTTGAACCATCTGAAAAACCGATCTTTAACGCCAAGACCATTGAACA ATCCCCATCATTCGGCAAATTTGAGATCGACACGGACGCAAACGTTCCAAGAGAT TTGTTTGAATATACTTTGGCTAACCAAATGTTGACAGCAATGGCTCAAGGTTATG CTGCTGAAATTTCCGCCAGAAGAAACGCTATGGATAACGCTTCCAAGAATGCCGG TGATATGATCAATCGTTACTCTATCTTGTACAACAGAACAAGACAAGCTGTCATT ACTAATGAACTGGTTGATATTATTACTGGTGCTTCCTCTTTGGGATGATAAAAAA ATCACCCTGCATTGCCTCTTTATTTGACGTTGTTTTTGTAGAACATGAAACGAAT TTTGACTTGATGAGACGAAGTATATATCCAAAGAATACCTTAAATAGAAAAGGAA AGATAATAAATACTAAACACTACTATATATTCAGGTAAAATACAAAAATTATAAC ATTTTTAAAACTTTTTTTTTTTGAAAGTCCTTCTCGCTTTAGGATTTTTTCCCAT TAAGATTATGATGGTTTCATGTATGTGTCAGTTAAAAAAAATATGGCTAAGTAGC AATGAAACGAACCAAGAAAATGAAAACAATATATAGTGCTGTTGAAATAACAAAG ACATTGGTATATATTTGTAAAATGTCTGTTAAATGTTTTTTATCTCAGGTTCTTG CTTGTCTTTGGTAGAAGAAATTATAGTAAACAAACAAACAAACAAACAAAAAAAA AAAAAAAAAA
pAdh
TTAAAACAAGAAGAGGGTTGACTACATCACGATGAGGGGGATCGAAGAAATGATG GTAAATGAAATAGGAAATCAAGGAGCATGAAGGCAAAAGACAAATATAAGGGTCG AACGAAAAATAAAGTGAAAAGTGTTGATATGATGTATTTGGCTTTGCGGCGCCGA AAAAACGAGTTTACGCAATTGCACAATCATGCTGACTCTGTGGCGGACCCGCGCT CTTGCCGGCCCGGCGATAACGCTGGGCGTGAGGCTGTGCCCGGCGGAGTTTTTTG CGCCTGCATTTTCCAAGGTTTACCCTGCGCTAAGGGGCGAGATTGGAGAAGCAAT AAGAATGCCGGTTGGGGTTGCGATGATGACGACCACGACAACTGGTGTCATTATT TAAGTTGCCGAAAGAACCTGAGTGCATTTGCAACATGAGTATACTAGAAGAATGA GCCAAGACTTGCGAGACGCGAGTTTGCCGGTGGTGCGAACAATAGAGCGACCATG ACCTTGAAGGTGAGACGCGCATAACCGCTAGAGTACTTTGAAGAGGAAACAGCAA TAGGGTTGCTACCAGTATAAATAGACAGGTACATACAACACTGGAAATGGTTGTC TGTTTGAGTACGCTTTCAATTCATTTGGGTGTGCACTTTATTATGTTACAATATG GAAGGGAACTTTACACTTCTCCTATGCACATATATTAATTAAAGTCCAATGCTAG TAGAGAAGGGGGGTAACACCCCTCCGCGCTCTTTTCCGATTTTTTTCTAAACCGT GGAATATTTCGGATATCCTTTTGTTGTTTCCGGGTGTACAATATGGACTTCCTCT TTTCTGGCAACCAAACCCATACATCGGGATTCCTATAATACCTTCGTTGGTCTCC CTAACATGTAGGTGGCGGAGGGGAGATATACAATAGAACAGATACCAGACAAGAC ATAATGGGCTAAACAAGACTACACCAATTACACTGCCTCATTGATGGTGGTACAT AACGAACTAATACTGTAGCCCTAGACTTGATAGCCATCATCATATCGAAGTTTCA CTACCCTTTTTCCATTTGCCATCTATTGAAGTAATAATAGGCGCATGCAACTTCT TTTCTTTTTTTTTCTTTTCTCTCTCCCCCGTTGTTGTCTCACCATATCCGCAATG ACAAAAAAATGATGGAAGACACTAAAGGAAAAAATTAACGACAAAGACAGCACCA ACAGATGTCGTTGTTCCAGAGCTGATGAGGGGTATCTCGAAGCACACGAAACTTT TTCCTTCCTTCATTCACGCACACTACTCTCTAATGAGCAACGGTATACGGCCTTC CTTCCAGTTACTTGAATTTGAAATAAAAAAAAGTTTGCTGTCTTGCTATCAAGTA TAAATAGACCTGCAATTATTAATCTTTTGTTTCCTCGTCATTGTTCTCGTTCCCT TTCTTCCTTGTTTCTTTTTCTGCACAATATTTCAAGCTATACCAAGCATACAATC AACTATCTCATATACA
pCyc
CAGATCCGCCAGGCGTGTATATATAGCGTGGATGGCCAGGCAACTTTAGTGCTGA CACATACAGGCATATATATATGTGTGCGACGACACATGATCATATGGCATGCATG TGCTCTGTATGTATATAAAACTCTTGTTTTCTTCTTTTCTCTAAATATTCTTTCC TTATACATTAGGACCTTTGCAGCATAAATTACTATACTTCTATAGACACACAAAC ACAAATACACACACTAAATTAATA
pSte5
CACGAAGTGACTGACAGAATACTGACATCAGCTGATTTCTCATAGAGCTGTTTCT CTGATAACACGTTGTTTGAACATCGACAAGATGAAAATCTAGAAGTATCAAGTTT CCTTTAAAGGGATATATAACAGATTCTAAAACTGACAGAAATATTTCGAGTGAAG AAGAAGCGTTAAATATTGGATCTTTCCGCAGTTCTACTCTGATACATTTTTGAAG TAGGAGAGTCATTTAGAAGGCGTATTGCTCAATAGTAGAAAGCAGGCCTGTGCAC ATGAATTAATTAAAAAATATAAAGGTAGTGATTAGACGACACATGTCCATAGGTA ACCTGTCATAATTTTGAACAATTTCCCTTCTTTTCTTTTTTTTTTTTGGGTGCGG CGATATGTAGCTTGTTAATTTACACATCATGTACTTTTCTGCATCAAAATATGAA AGGCGATAGTAGCTAAAGAAAATACCGAGAATTTCCTCGAAAAGTTGACGACAAA AGAAAGGCATAAAAAAGTAATTTGAAAATATTTTAAAACTGTTTTAACCCATCTA GCATCCGCGCTAAAAAAGGAAGATACAGGATACAGCGGAAACAACTTTTAA
2-ATG-XhoI-“A” and 2 adaptor bases:
ATGCTCGAGGGAGCT
3-mCherry:
GTGAGCAAGGGCGAGGAGGATAACATGGCCATCATCAAGGAGTTCATGCGCTTCAAGGTGCACATGGAGGGCTCCGTGAACGGCCACG AGTTCGAGATCGAGGGCGAGGGCGAGGGCCGCCCCTACGAGGGCACCCAGACCGCCAAGCTGAAGGTGACCAAGGGTGGCCCCCTGCC CTTCGCCTGGGACATCCTGTCCCCTCAGTTCATGTACGGCTCCAAGGCCTACGTGAAGCACCCCGCCGACATCCCCGACTACTTGAAG CTGTCCTTCCCCGAGGGCTTCAAGTGGGAGCGCGTGATGAACTTCGAGGACGGCGGCGTGGTGACCGTGACCCAGGACTCCTCCCTGC AGGACGGCGAGTTCATCTACAAGGTGAAGCTGCGCGGCACCAACTTCCCCTCCGACGGCCCCGTAATGCAGAAGAAGACCATGGGCTG GGAGGCCTCCTCCGAGCGGATGTACCCCGAGGACGGCGCCCTGAAGGGCGAGATCAAGCAGAGGCTGAAGCTGAAGGACGGCGGCCAC TACGACGCTGAGGTCAAGACCACCTACAAGGCCAAGAAGCCCGTGCAGCTGCCCGGCGCCTACAACGTCAACATCAAGTTGGACATCA CCTCCCACAACGAGGACTACACCATCGTGGAACAGTACGAACGCGCCGAGGGCCGCCACTCCACCGGCGGCATGGACGAGCTGTACAAG
4-Adaptor bases, followed by “B” and another adaptor base: GGTAGTTCCCTA
5-Effectors
For YopH:
AACTTATCATTAAGCGATCTTCATCGTCAGGTATCTCGATTGGTGCAGCAAGAGAGCGGTGATTGTACCGGGAAATTAAGAGGTAACGT TGCTGCCAATAAAGAAACTACCTTTCAAGGTTTGACCATAGCCAGTGGAGCCAGAGAGTCAGAAAAAGTATTTGCTCAAACTGTACTAA GCCACGTAGCAAATGTTGTTCTAACTCAAGAAGATACCGCTAAGCTATTGCAAAGTACGGTAAAGCATAATTTGAATAATTATGACTTA AGAAGTGTCGGCAATGGTAATAGTGTACTTGTCAGTTTACGTAGTGACCAAATGACACTACAAGACGCCAAAGTGCTGTTGGAGGCCGC ATTGCGACAAGAGTCGGGAGCGAGGGGGCATGTATCATCTCATTCACATTCAGCCCTTCACGCACCGGGAACCCCGGTGCGTGAAGGAC TGCGTTCACATCTAGACCCCAGAACTCCACCGTTGCCACCGCGTGAACGACCACACACTTCTGGCCATCACGGGGCTGGCGAAGCCAGA GCCACCGCACCAAGCACTGTTTCTCCTTATGGCCCAGAAGCGCGCGCAGAACTCAGCAGCCGCCTCACCACATTGCGCAATACGCTGGC GCCAGCAACGAATGATCCGCGTTACTTACAAGCCTGCGGCGGTGAAAAGCTAAACCGATTTAGAGATATTCAATGCTGTCGGCAAACCG CAGTACGCGCCGATCTTAATGCCAATTACATCCAGGTCGGTAACACTCGTACCATAGCGTGCCAGTATCCGCTACAATCTCAACTTGAA AGCCATTTCCGTATGCTGGCAGAAAACCGAACGCCAGTGTTGGCTGTTTTAGCGTCCAGTTCTGAGATAGCCAATCAAAGATTCGGTAT GCCAGATTATTTCCGCCAGAGTGGTACCTATGGCAGTATCACTGTAGAGTCTAAAATGACTCAGCAAGTTGGTCTCGGTGACGGGATTA TGGCAGATATGTATACTTTAACGATTCGTGAAGCGGGTCAAAAAACAATCTCTGTTCCTGTGGTTCATGTTGGCAATTGGCCCGATCAG ACCGCAGTCAGCTCTGAAGTTACCAAGGCACTCGCTTCACTGGTAGATCAAACAGCAGAAACAAAACGCAATATGTATGAAAGCAAAGG AAGTTCAGCGGTAGGAGATGACTCCAAATTACGGCCGGTAATACATTGCCGTGCGGGTGTTGGCCGTACTGCGCAACTGATTGGCGCAA TGTGCATGAATGATAGTCGTAATAGTCAGTTAAGCGTAGAAGATATGGTCAGCCAAATGCGAGTACAAAGAAATGGTATTATGGTACAA AAAGATGAGCAACTTGATGTTCTGATTAAGTTGGCTGAAGGACAAGGGCGACCATTATTAAATAGC
For YopJ:
ATCGGACCAATATCACAAATAAATATCTCCGGTGGCTTATCAGAAAAAGAGACCAGTTCTTTAATCAGTAATGAAGAGCTTAAAAATAT CATAACACAGTTGGAAACTGATATATCGGATGGATCCTGGTTCCATAAAAATTATTCACGTATGGATGTAGAAGTCATGCCCGCATTGG TAATCCAGGCGAACAATAAATATCCGGAAATGAATCTTAATCTTGTTACATCTCCATTGGACCTTTCAATAGAAATAAAAAACGTCATA GAAAATGGAGTTAGATCTTCCCGCTTCATAATTAACATGGGGGAAGGTGGAATACATTTCAGTGTAATTGATTACAAACATATAAATGG GAAAACATCTCTGATATTGTTTGAACCAGCAAACTTTAACAGTATGGGGCCAGCGATGCTGGCAATAAGGACAAAAACGGCTATTGAAC GTTATCAATTACCTGATTGCCATTTCTCCATGGTGGAAATGGATATTCAGCGAAGCTCATCTGAATGTGGTATTTTTAGTTTGGCACTG GCAAAAAAACTTTACATCGAGAGAGATAGCCTGTTGAAAATACATGAAGATAATATAAAAGGTATATTAAGTGATGGTGAAAATCCTTT ACCCCACGATAAGTTGGACCCGTATCTCCCGGTAACTTTTTACAAACATACTCAAGGTAAAAAACGTCTTAATGAATATTTAAATACTA ACCCGCAGGGAGTTGGTACTGTTGTTAACAAAAAAAATGAAACCATCGTTAATAGATTTGATAACAATAAATCCATTGTAGATGGAAAG GATTATCAGTTTCGGTACATAAAAAGAGAATAGCTGAATATAAAACACTTCTCAAAGTA
For OspF:
CCAATTAAAAAGCCTTGTTTGAAATTGAATTTAGATTCTTTGAACGTTGTTAGATCAGAAATTCCACAAATGTTATCTGCTAATGAAAGAT TGAAAAATAATTTTAACATTTTATATAATCAAATAAGACAGTACCCAGCTTATTATTTTAAAGTTGCTTCTAATGTTCCTACTTACTCTGA TATATGTCAATTTTTCTCAGTTATGTATCAAGGTTTTCAGATAGTTAATCATTCTGGTGATGTTTTTATTCATGCTTGTAGAGAAAATCCA CAATCTAAAGGTGATTTTGTTGGTGACAAGTTCCATATTTCTATAGCTAGAGAACAAGTTCCATTGGCATTTCAAATTTTATCTGGTTTGTT ATTTTCTGAGGACTCTCCTATTGACAAATGGAAGATAACTGATATGAATAGAGTTTCTCAACAGTCAAGAGTTGGTATTGGTGCTCAATTTAC TTTGTATGTTAAATCTGATCAAGAATGTTCTCAGTATTCAGCTTTATTGTTGCATAAAATTAGACAATTCATAATGTGCTTGGAATCTAATTT GTTAAGATCTAAGATTGCTCCAGGTGAATATCCTGCATCTGATGTTAGACCAGAGGATTGGAAGTACGTTTCATATAGAAATGAATTGAGATC TGACAGAGATGGTTCTGAAAGACAAGAGCAGATGTTAAGAGAAGAACCTTTTTATAGATTGATGATTGAG
6-Adaptor bases, followed by “C” followed by another adaptor: GGTAGCGAT
7-Zippers: For RR: GGTTCTGGTTCTGGTTCTGGTTCTCTGGAAATCCGTGCGGCGTTCCTGGAAAAAGAAAACACCGCGCTGCGTACCCGTGCGGCGGAACTGCG TAAACGTGTTGGTCGTTGCCGTAACATCGTTTCTAAATACGAAACCCGTTACGGTCCGCTG For EE:
GGTTCCGGTAGCGGTTCCGGTTCTATCACTATTAGGGCTGCTTTTTTGGAAAAAGAAAATACTGCTTTGAGAACTGAAATTGCTGAATTGG AAAAAGAAGTTGGTAGATGTGAAAATATTGTTTCTAAATATGAAACTAGATATGGTCCATTG
8-Final adaptors, followed by “D”, followed by BamHI, 3 STOPS in three different frames, then NotI, the Adh terminator and SacI:
TAATGCGGGATCCTGAGTAAATAAGCGGCCGCGCGAATTTC
TTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTAT
ACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGT
AACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT
ATTGACCACACCGAGCTC
PROJECT 2
(1) PspOMI//pSte2-Ste2//XhoI//GFP//XmaI//RR//STOP//BamHI//AdhTerminator
GGGCCCATCCAATATCACCTGACCTTCATCTTTATGTTTTTCCACTGGTGTAATGTACTCTGTTGCATCATCAATAAGCAACTGGGACGCCAATAATTCATACCAACAATCTTTTCTTAATGAGTTGTTTACAAACCCCAGAGGCCCTAAACCTATGTACCTCAAAGCTCGGACATCTTTTTTCTCTAATAACTTTGATATAACTTTTCCTTTTTGTTCTTCTGCCTTTCCACTGTCACAACGAGAAAGTGATTCCAGATATGCGTTATAACCTCCCCTTGTCAACGCGTCTTTGTCGTGGCTACTCTGATTAGTATGAAATAATGACCTTAATGGCATCGTATTTTGTTAATTGGCTTGTTCTATGTCATTTGCTAAATTTTCTCTTTATTTCTTTACTTTGCCAATTCAAGATAATCTTCCCTTCCCAGAGAGAAAAAAGGAAAAATTTAGCTATGAAACCTCAATAAGCTTTTTAATACACCAAAGATTCAAGATAAGAGCATAGAACGAACTGTAGAATAGTCCGGATATGTTATCCAATGCCTGCCAAAATGCATTGTCACACGCTGTAGTGCTCGAATAGGTGTTGCAATCCGTCAATATACGTCTTGCTCTGTGGGTAAATGTCTCGTGCATTAAGACAGGCTAGTATAAACGAGAAGAAGTATCCTGCTTTGCAATGAAACAATAGTATCCGCTAAGAATTTAAGCAGGCCAACGTCCATACTGCTTAGGACCTGTGCCTGGCAAGTCGCAGATTGAAGTTTTTTCAACCATGTAAATTTCCTAATTGGGTAAGTACATGATGAAACACATATGAAGAAAAAAGCTTTCCTACATATTCAAGATTTTTTTCTGTGGGTGGAATACTATTTAAGGAGTGCTATTAGTATCTTATTTGACTTCAAAGCAATACGATACCTTTTCTTTTCACCTGCTCTGGCTATAATTATAATTGGTTACTTAAAAATGCACCGTTAAGAACCATATCCAAGAATCAAAAATGTCTGATGCGGCTCCTTCATTGAGCAATCTATTTTATGATCCAACGTATAATCCTGGTCAAAGCACCATTAACTACACTTCCATATATGGGAATGGATCTACCATCACTTTCGATGAGTTGCAAGGTTTAGTTAACAGTACTGTTACTCAGGCCATTATGTTTGGTGTCAGATGTGGTGCAGCTGCTTTGACTTTGATTGTCATGTGGATGACATCGAGAAGCAGAAAAACGCCGATTTTCATTATCAACCAAGTTTCATTGTTTTTAATCATTTTGCATTCTGCACTCTATTTTAAATATTTACTGTCTAATTACTCTTCAGTGACTTACGCTCTCACCGGATTTCCTCAGTTCATCAGTAGAGGTGACGTTCATGTTTATGGTGCTACAAATATAATTCAAGTCCTTCTTGTGGCTTCTATTGAGACTTCACTGGTGTTTCAGATAAAAGTTATTTTCACAGGCGACAACTTCAAAAGGATAGGTTTGATGCTGACGTCGATATCTTTCACTTTAGGGATTGCTACAGTTACCATGTATTTTGTAAGCGCTGTTAAAGGTATGATTGTGACTTATAATGATGTTAGTGCCACCCAAGATAAATACTTCAATGCATCCACAATTTTACTTGCATCCTCAATAAACTTTATGTCATTTGTCCTGGTAGTTAAATTGATTTTAGCTATTAGATCAAGAAGATTCCTTGGTCTCAAGCAGTTCGATAGTTTCCATATTTTACTCATAATGTCATGTCAATCTTTGTTGGTTCCATCGATAATATTCATCCTCGCATACAGTTTGGAACCAAACCAGGGAACAGATGTCTTGACTACTGTTGCAACATTACTTGCTGTATTGTCTTTACCATTATCATCAATGTGGGCCACGGCTGCTAATAATGCATCCAAAACAAACACAATTACTTCAGACTTTACAACATCCACAGATAGGTTTTATCCAGGCACGCTGTCTAGCTTTCAAACTGATAGTATCAACAACGATGCTAAAAGCAGTCTCAGAAGTAGATTATATGACCTATATCCTAGAAGGAAGGAAACAACATCGGATAAACATTCGGAAAGAACTTTTGTTTCTGAGACTGCAGATGATATAGAGAAAAATCAGTTTTATCAGTTGCCCACACCTACGAGTTCAAAAAATACTAGGATAGGACCGTTTGCTGATGCAAGTTACAAAGAGGGAGAAGTTGAACCCGTCGACATGTACACTCCCGATACGGCAGCTGATGAGGAAGCCAGAAAGTTCTGGACTGAAGATAATAATAATTTACTCGAGTCTAAAGGTGAAGAATTATTCACTGGTGTTGTCCCAATTTTGGTTGAATTAGATGGTGATGTTAATGGTCACAAATTTTCTGTCTCCGGTGAAGGTGAAGGTGATGCTACTTACGGTAAATTGACCTTAAAATTTATTTGTACTACTGGTAAATTGCCAGTTCCATGGCCAACCTTAGTCACTACTTTAACTTATGGTGTTCAATGTTTTTCTAGATACCCAGATCATATGAAACAACATGACTTTTTCAAGTCTGCCATGCCAGAAGGTTATGTTCAAGAAAGAACTATTTTTTTCAAAGATGACGGTAACTACAAGACCAGAGCTGAAGTCAAGTTTGAAGGTGATACCTTAGTTAATAGAATCGAATTAAAAGGTATTGATTTTAAAGAAGATGGTAACATTTTAGGTCACAAATTGGAATACAACTATAACTCTCACAATGTTTACATCATGGCTGACAAACAAAAGAATGGTATCAAAGTTAACTTCAAAATTAGACACAACATTGAAGATGGTTCTGTTCAATTAGCTGACCATTATCAACAAAATACTCCAATTGGTGATGGTCCAGTCTTGTTACCAGACAACCATTACTTATCCACTCAATCTaggTTATCCAAAGATCCAAACGAAAAGAGAGACCACATGGTCTTGTTAGAATTTGTTACTGCTGCTGGTATTACCCATGGTATGGATGAATTGTACAAACCCGGGggatctggatcgggatctGATCCAGATTTGGAAATTAGAGCTGCTTTTTTGAGACAAAGAAATACTGCTTTGAGAACTGAAGTTGCTGAATTGGAACAAGAAGTTCAAAGATTGGAAAATGAAGTTTCTCAATATGAAACTAGATATGGTCCATTGGGTGGTGGTAAATAAGGATCCGCGAATTTCTTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTATACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGTAACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT ATTGACCACACC
(2) PspOMI//pGal1//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator
TGTTATTGTGATAGTATCTCAGCGGGATTTGTGTAGATTCACCTACAGCCTTTAAACCAGCAAAGTGAATTACCGAATCAATTTTATATTCTTTGAAAACCTTTTCCAGACCTTTTCGGTCACACAAATCAACCTCATAGAAGGGAATGTGATGCTTGGTCAAGACCTCTAACCTGGCTACAGAATCATAAGTTGAATTCGACAGGTTATCAGCAACAACACAGTCATATCCATTCTCAATTAGCTCTACCACAGTGTGTGAACCAATGTATCCAGCACCACCTGTAACCAAAACAATTTTAGAAGTACTTTCACTTTGTAACTGAGCTGTCATTTATATTGAATTTTCAAAAATTCTTACTTTTTTTTTGGATGGACGCAAAGAAGTTTAATAATCATATTACATGGCATTACCACCATATACATATCCATATCTAATCTTACTTATATGTTGTGGAAATGTAAAGAGCCCCATTATCTTAGCCTAAAAAAACCTTCTCTTTGGAACTTTCAGTAATACGCTTAACTGCTCATTGCTATATTGAAGTACGGATTAGAAGCCGCCGAGCGGGCGACAGCCCTCCGACGGAAGACTCTCCTCCGTGCGTCCTCGTCTTCACCGGTCGCGTTCCTGAAACGCAGATGTGCCTCGCGCCGCACTGCTCCGAACAATAAAGATTCTACAATACTAGCTTTTATGGTTATGAAGAGGAAAAATTGGCAGTAACCTGGCCCCACAAACCTTCAAATTAACGAATCAAATTAACAACCATAGGATGATAATGCGATTAGTTTTTTAGCCTTATTTCTGGGGTAATTAATCAGCGAAGCGATGATTTTTGATCTATTAACAGATATATAAATGGAAAAGCTGCATAACCACTTTAACTAATACTTTCAACATTTTCAGTTTGTATTACTTCTTATTCAAATGTCATAAAAGTATCAACAAAAAATTGTTAATATACCTCTATACTTTAACGTCAAGGAGAAAAAACTATAATGCTCGAGGGAGCTgcttctgcatcaacttctaaatataattcacactccttggagaatgagtctattaagaggacgtctcgagatggagtcaatcgagatctcactgaggctgttcctcgacttccaggagaaacactaatcactgacaaagaagttatttacatatgtcctttcaatggccccattaagggaagagtttacatcacaaattatcgtctttatttaagaagtttggaaacggattcttctctaatacttgatgttcctctgggtgtgatctcgagaattgaaaaaatgggaggcgcgacaagtagaggagaaaattcctatggtctagatattacttgtaaagacatgagaaacctgaggttcgctttgaaacaggaaggccacagcagaagagatatgtttgagatcctcacgagatacgcgtttcccctggctcacagtctgccattatttgcatttttaaatgaagaaaagtttaacgtggatggatggacagtttacaatccagtggaagaatacaggaggcagggcttgcccaatcaccattggagaataacttttattaataagtgctatgagctctgtgacacttaccctgctcttttggtggttccgtatcgtgcctcagatgatgacctccggagagttgcaacttttaggtcccgaaatcgaattccagtgctgtcatggattcatccagaaaataagacggtcattgtgcgttgcagtcagcctcttgtcggtatgagtgggaaacgaaataaagatgatgagaaatatctcgatgttatcagggagactaataaacaaatttctaaactcaccatttatgatgcaagacccagcgtaaatgcagtggccaacaaggcaacaggaggaggatatgaaagtgatgatgcatatcataacgccgaacttttcttcttagacattcataatattcatgttatgcgggaatctttaaaaaaagtgaaggacattgtttatcctaatgtagaagaatctcattggttgtccagtttggagtctactcattggttagaacatatcaagctcgttttgacaggagccattcaagtagcagacaaagtttcttcagggaagagttcagtgcttgtgcattgcagtgacggatgggacaggactgctcagctgacatccttggccatgctgatgttggatagcttctataggagcattgaagggttcgaaatactggtacaaaaagaatggataagttttggacataaatttgcatctcgaataggtcatggtgataaaaaccacaccgatgctgaccgttctcctatttttctccagtttattgattgtgtgtggcaaatgtcaaaacagttccctacagcttttgaattcaatgaacaatttttgattataattttggatcatctgtatagttgccgatttggtactttcttattcaactgtgaatctgctcgagaaagacagaaggttacagaaaggactgtttctttatggtcactgataaacagtaataaagaaaaattcaaaaaccccttctatactaaagaaatcaatcgagttttatatccagttgccagtatgcgtcacttggaactctgggtgaattactacattagatggaaccccaggatcaagcaacaacagccgaatccagtggagcagcgttacatggagctcttagccttacgcgacgaatacataaagcggcttgaggaactgcagctcgccaactctgccaagctttctgatcccccaacttcaccttccagtccttcgcaaatgatgccccatgtgcaaactcacttcGGTAGTTCCCTAggttccggtagcggttccggttctatcactattagggctgcttttttggaaaaagaaaatactgctttgagaactgaaattgctgaattggaaaaagaagttggtagatgtgaaaatattgtttctaaatatgaaactagatatggtccattgTAATCGCGGATCCGCGAATTTCTTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTATACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGTAACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT ATTGACCACACC
(3) pGal1//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator
As above, but WMTM1 sequence
(4) pSte5//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator
As above, but Ste5 promoter sequence:
CACGAAGTGACTGACAGAATACTGACATCAGCTGATTTCTCATAGAGCTGTTTCTCTGATAACACGTTGTTTGAACATCGACAAGATGAAAATCTAGAAGTATCAAGTTTCCTTTAAAGGGATATATAACAGATTCTAAAACTGACAGAAATATTTCGAGTGAAGAAGAAGCGTTAAATATTGGATCTTTCCGCAGTTCTACTCTGATACATTTTTGAAGTAGGAGAGTCATTTAGAAGGCGTATTGCTCAATAGTAGAAAGCAGGCCTGTGCACATGAATTAATTAAAAAATATAAAGGTAGTGATTAGACGACACATGTCCATAGGTAACCTGTCATAATTTTGAACAATTTCCCTTCTTTTCTTTTTTTTTTTTGGGTGCGGCGATATGTAGCTTGTTAATTTACACATCATGTACTTTTCTGCATCAAAATATGAAAGGCGATAGTAGCTAAAGAAAATACCGAGAATTTCCTCGAAAAGTTGACGACAAAAGAAAGGCATAAAAAAGTAATTTGAAAATATTTTAAAACTGTTTTAACCCATCTAGCATCCGCGCTAAAAAAGGAAGATACAGGATACAGCGGAAACAACTTTTAAATG
(5) pSte5//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator
As above.
(6) pCyc//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator
As above, but Cyc promoter sequence:
ctcgagcagatccgccaggcgtgtatatatagcgtggatggccaggcaactttagtgctgacacatacaggcatatatatatgtgtgcgacgacacatgatcatatggcatgcatgtgctctgtatgtatataaaactcttgttttcttcttttctctaaatattctttccttatacattaggacctttgcagcataaattactatacttctatagacacacaaacacaaatacacacactaaattaataatg
(7) pCyc//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator
As above.
(8) pCyc//ATG//XhoI//GFP//XmaI//(SerGlySerGlySer)ING2 PH domain//STOP//BamHI
As above, but //(SerGlySerGlySer)ING2 PH domain sequence as follows:
TCTGGATCAGGTTCAGCTTCTCCAGTTGAATTTGCTATTGATCCTAACGAACCTACATACTGCTTATGTAATCAAGTTTCTTATGGTGAAATGATTGGTTGCGATAACGAGCAGTGTCCAATAGAATGGTTTCATTTCTCTTGCGTTTCTTTAACATATAAACCTAAGGGAAAATGGTACTGTCCTAAATGTAGAGGTGATAATGAGAAAACTATGGATAAGTCTACTGAAAAGACAAAAAAAGATAGAAGATCAAGA
