LOCUS       sequence-254101-        7049 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast pathway position 5. PYC transcription unit with the HHF2 
            promoter and ADH1 terminator..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7049)
  AUTHORS   Young EM, Zhao Z, Gielesen BEM, Wu L, Benjamin Gordon D, Roubos JA, 
            Voigt CA
  TITLE     Iterative algorithm-guided design of massive strain libraries, 
            applied to itaconic acid production in yeast.
  JOURNAL   Metab Eng. 2018 Jul;48:33-43. doi: 10.1016/j.ymben.2018.05.002. Epub
            2018 May 9.
  PUBMED    29753070
REFERENCE   2  (bases 1 to 7049)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Metab 
            Eng."; date: "2018-07"; volume: "48:33-43. doi"; pages: " 
            10.1016/j.ymben.2018.05.002. Epub 2018 May 9"
COMMENT     Sequence Label: pDSM-de-Phhf2-PYC-Tadh1
FEATURES             Location/Qualifiers
     source          1..7049
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(3965..3998)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(4468..4485)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4468..4484)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4583..4600)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4754..5342)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4834..4853)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5464..6273)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     promoter        complement(6274..6365)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6432..6451
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     protein_bind    6658..6691
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
ORIGIN
        1 atgcagacca gaccagaaca aaacaactga caagaaggcg tttaatttaa tatgttgttc
       61 actcgcgcct gggctgttgt tattcggcta gatacatacg tgtttgtgcg tatgtagtta
      121 tatcatatat aagtatatta ggatgaggcg gtgaaagaga tttttttttt ttcgcttaat
      181 ttattctttt ctctatcttt tttcctacat cttgttcaaa agagtagcaa aaacaacaat
      241 caatacaata aaataatgtc ctcttccaag atcttggctg gtttgagaga caacttttct
      301 ttgttgggtg aaaagaacaa gattttggtc gccaacagag gtgaaatccc aatcagaatt
      361 ttcagatctg ctcacgaatt gtctatgaga actatcgcca tctactctca cgaagataga
      421 ttatccatgc acagattgaa ggctgatgaa gcctacgtta tcggtgaaga aggtcaatac
      481 accccagtcg gtgcttactt ggccatggac gaaatcatcg aaattgccaa gaagcacaag
      541 gtcgatttca tccacccagg ttacggtttc ttgtctgaaa actctgaatt tgctgacaag
      601 gttgttaagg ctggtattac ctggattggt ccaccagctg aagtcattga atctgttggt
      661 gacaaggttt ctgccagaca tttggctgct cgtgccaacg ttccaactgt cccaggtact
      721 ccaggtccta tcgaaaccgt tcaagaagct ctagatttcg tcaatgaata cggttaccca
      781 gttatcatca aggctgcttt cggtggtggt ggtcgtggta tgagagttgt cagagaaggt
      841 gacgatgtcg ctgatgcttt ccaaagagcc acttctgaag ctagaactgc tttcggtaac
      901 ggtacttgtt tcgtcgaaag attcttggac aagccaaagc acattgaagt tcaattatta
      961 gctgacaacc acggtaacgt tgtccacttg ttcgaaagag actgttccgt ccaaagacgt
     1021 caccaaaagg ttgtcgaagt tgctccagct aagactttac caagagaagt tagagatgct
     1081 atcttgaccg atgccgttaa gttggctaag gtttgtggtt acagaaacgc tggtactgct
     1141 gaattcttgg ttgacaacca aaacagacat tacttcattg aaatcaaccc aagaattcaa
     1201 gtcgaacaca ccatcactga agaaatcact ggtattgaca ttgtctccgc tcaaatccaa
     1261 atcgccgctg gtgctacttt gactcaatta ggtctattac aagacaaaat caccaccaga
     1321 ggtttctcta tccaatgtcg tatcaccact gaagatccat ccaagaactt ccaaccagac
     1381 actggtcgtt tggaagtcta cagatccgct ggtggtaacg gtgtcagatt ggacggtggt
     1441 aacgcctacg ctggtgctac catctctcca cactacgact ccatgttggt taagtgttcc
     1501 tgttctggtt ctacctacga aattgtcaga agaaagatga tcagagcttt gattgaattc
     1561 agaatcagag gtgtcaagac caacatccca ttcttgttga ctttgttgac caacccagtt
     1621 ttcattgaag gtacctactg gaccactttc atcgatgaca ctccacaatt gttccaaatg
     1681 gtttcctctc aaaacagagc tcaaaaattg ttgcactact tggctgactt ggccgtcaac
     1741 ggttcctcta tcaagggtca aatcggttta ccaaagttga agtccaaccc ttccgttcca
     1801 catttgcacg atgctcaagg taatgtcatc aacgttacca aatctgcccc accatccggt
     1861 tggagacaag tcttgttgga aaagggtcca tccgaatttg ccaagcaagt cagacaattc
     1921 aacggtactt tgttgatgga caccacctgg agagatgctc accaatcttt gctagctacc
     1981 agagtcagaa ctcacgattt ggccaccatt gctccaacca ctgctcacgc tttggctggt
     2041 gcctttgctt tggaatgttg gggtggtgct actttcgatg tcgccatgag attcttgcat
     2101 gaggacccat gggaaagatt gagaaaattg agatctttgg tcccaaacat tccattccaa
     2161 atgttgttga gaggtgctaa cggtgttgct tactcctctt tgccagacaa cgccattgac
     2221 catttcgtta agcaagccaa ggacaatggt gttgacattt tcagagtctt tgacgctttg
     2281 aacgacttgg aacaattgaa ggttggtgtt aatgctgtca agaaggctgg tggtgttgtc
     2341 gaagctaccg tttgttactc tggtgacatg ttgcaaccag gtaagaaata caacttggac
     2401 tactacttag aagttgtcga aaagatcgtt caaatgggta ctcacatctt gggtatcaag
     2461 gacatggctg gtaccatgaa gccagctgct gccaaattgt tgattggttc tttacgtacc
     2521 agatacccag acttgccaat ccacgttcac tctcatgact ccgctggtac tgctgttgct
     2581 tccatgactg cttgtgcttt ggccggtgct gatgttgttg acgttgccat taactccatg
     2641 tccggtttga cctctcaacc atctattaac gctttgttgg cctccttgga aggtaacatt
     2701 gacactggta tcaacgtcga acacgttaga gaattggacg cttactgggc tgaaatgaga
     2761 ttattatact cttgtttcga agctgacttg aagggtccag accctgaagt ttaccaacac
     2821 gaaattccag gtggtcaatt gaccaacttg ttgttccaag ctcaacaatt aggtctaggt
     2881 gaacaatggg ctgaaaccaa gagagcttac agagaagcta actacttgtt gggtgacatt
     2941 gttaaggtca ccccaacttc taaggtcgtt ggtgatttgg ctcaattcat ggtttctaac
     3001 aaattgactt ctgatgacat cagaagatta gctaactctt tggacttccc agactccgtt
     3061 atggacttct tcgaaggttt gatcggtcaa ccatacggtg gtttcccaga accattgaga
     3121 tccgatgttt tgagaaacaa gcgtcgtaaa ttgacttgta gaccaggttt agaattggaa
     3181 ccattcgatt tggaaaagat cagagaagat ttgcaaaaca gattcggtga tatcgatgaa
     3241 tgtgatgttg cctcctacaa catgtatcct cgtgtctacg aagatttcca aaagattaga
     3301 gaaacttacg gtgacttgtc tgtcttacca accaagaact tcttggctcc agctgaacca
     3361 gacgaagaaa tcgaagtcac cattgaacaa ggtaagactt tgattatcaa attacaagct
     3421 gttggtgatt tgaacaagaa aaccggtcaa agagaagtct acttcgaatt gaacggtgaa
     3481 ttgagaaaga tcagagttgc tgacaaatct caaaacattc aatctgttgc caagccaaag
     3541 gctgatgtcc acgacaccca ccaaatcggt gctccaatgg ctggtgtcat cattgaagtc
     3601 aaggttcaca agggttcttt ggtcaagaag ggtgaatcta tcgccgtttt gtctgctatg
     3661 aagatggaaa tggttgtttc ctctccagct gatggtcaag tcaaagatgt ctttatccgt
     3721 gacggtgaat ccgtcgatgc ttctgacttg ttggttgttt tggaagaaga aactctacca
     3781 ccttctcaaa agaaataaac gaatttctta tgatttatga tttttattat taaataagtt
     3841 ataaaaaaaa taagtgtata caaattttaa agtgacactt aggttttaaa acgaaaattc
     3901 cctcaaataa ccacaaacat ccttcccata tgctcggtcg tgcttgttgt acctaaaatg
     3961 aagtgaagtt cctatacttt ctagagaata ggaacttcta tagtgagtcg aataagggcg
     4021 acacaaaatt tattctaaat gcataataaa tactgataac atcttatagt ttgtattata
     4081 ttttgtatta tcgttgacat gtataatttt gatatcaaaa actgattttc cctttattat
     4141 tttcgagatt tattttctta attctcttta acaaactaga aatattgtat atacaaaaaa
     4201 tcataaataa tagatgaata gtttaattat aggtgttcat caatcgaaaa agcaacgtat
     4261 cttatttaaa gtgcgttgct tttttctcat ttataaggtt aaataattct catatatcaa
     4321 gcaaagtgac aggcgccctt aaatattctg acaaatgctc tttccctaaa ctccccccat
     4381 aaaaaaaccc gccgaagcgg gtttttacgt tatttgcgga ttaacgatta ctcgttatca
     4441 gaaccgccca gggggcccga gcttaagact ggccgtcgtt ttacaacaca gaaagagttt
     4501 gtagaaacgc aaaaaggcca tccgtcaggg gccttctgct tagtttgatg cctggcagtt
     4561 ccctactctc gccttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     4621 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     4681 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     4741 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     4801 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     4861 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     4921 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4981 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     5041 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     5101 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     5161 tgaagtggtg ggctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     5221 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     5281 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     5341 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgac gcgcgcgtaa
     5401 ctcacgttaa gggattttgg tcatgagctt gcgccgtccc gtcaagtcag cgtaatgctc
     5461 tgcttagaaa aactcatcga gcatcaaatg aaattgcaat ttattcatat caggattatc
     5521 aataccatat ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt
     5581 ccataggatg gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca
     5641 acctattaat ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac
     5701 gactgaatcc ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg
     5761 ccagccatta cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga
     5821 ttgcgcctga gcgaggcgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat
     5881 cgagtgcaac cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg
     5941 atattcttct aatacctgga acgctgtttt tccggggatc gcagtggtga gtaaccatgc
     6001 atcatcagga gtacggataa aatgcttgat ggtcggaagt ggcataaatt ccgtcagcca
     6061 gtttagtctg accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag
     6121 aaacaactct ggcgcatcgg gcttcccata caagcgatag attgtcgcac ctgattgccc
     6181 gacattatcg cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg
     6241 cggcctcgac gtttcccgtt gaatatggct catattcttc ctttttcaat attattgaag
     6301 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6361 acaaataggg gtcagtgtta caaccaatta accaattctg aacattatcg cgagcccatt
     6421 tatacctgaa tatggctcat aacacccctt gtttgcctgg cggcagtagc gcggtggtcc
     6481 cacctgaccc catgccgaac tcagaagtga aacgccgtag cgccgatggt agtgtgggga
     6541 ctccccatgc gagagtaggg aactgccagg catcaaataa aacgaaaggc tcagtcgaaa
     6601 gactgggcct ttcgcccggg ctaattaggg ggtgtcgccc ttattcgact ctatagtgaa
     6661 gttcctattc tctagaaagt ataggaactt ctgaagtggg gaacgttgtc caggtttgta
     6721 tccacgtgtg tccgttccgc caatattccg cgtgcacaag aagcaacgcg agagagcaca
     6781 acacgctgtt atcacgcaaa ctatgttttg acaccgagcc atagccgtga ttgtgcgtca
     6841 cattgggcga taatgaacgc taaatgacca actcccatcc gtaggagccc cttagggcgt
     6901 gccaatagtt tcacgcgctt aatgcgaagt gctcggaacg gacaactgtg gtcgtttggc
     6961 accgggaaag tggtactaga ccgagagttt cgcatttgta tggcaggacg ttctgggagc
     7021 ttcgcgtctc aagctttttc gggcgcgaa
//
