LOCUS       sequence-360449-        9067 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast integrative plasmid for expressing ERG20 (GAL10 promoter) and 
            fusion protein GFP-AcNES1 (GAL7 promoter). Contains uracil 
            auxotrophic marker (K. lactis URA3)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9067)
  AUTHORS   Cheah LC, Liu L, Stark T, Plan MR, Peng B, Lu Z, Schenk G, Sainsbury
            F, Vickers CE
  TITLE     Metabolic flux enhancement from the translational fusion of terpene 
            synthases is linked to terpene synthase accumulation.
  JOURNAL   Metab Eng. 2023 May;77:143-151. doi: 10.1016/j.ymben.2023.03.012. 
            Epub 2023 Mar 28.
  PUBMED    36990382
REFERENCE   2  (bases 1 to 9067)
  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: "2023-05"; volume: "77:143-151. doi"; pages: " 
            10.1016/j.ymben.2023.03.012. Epub 2023 Mar 28"
COMMENT     Sequence Label: Coexpressed Free GFP-NES
FEATURES             Location/Qualifiers
     source          1..9067
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    310..427
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     CDS             2460..3170
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(6190..6207)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6190..6206)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6199..6221)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(6415..6434)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6534..6556
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6594..6612)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6680..6784
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6785..6853,6854..7645)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(7003..7022)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7816..8404
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8305..8324
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8558..8575
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8692..8713
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8728..8733,8734..8751,8752..8758)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8766..8782
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     8771..8793
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8790..8806
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8790..8806
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        complement(join(9053..9067,1..427))
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     primer_bind     complement(join(9059..9067,1..15))
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
ORIGIN
        1 gtatagaggt atattaacaa ttttttgttg atacttttat gacatttgaa taagaagtaa
       61 tacaaaccga aaatgttgaa agtattagtt aaagtggtta tgcagctttt gcatttatat
      121 atctgttaat agatcaaaaa tcatcgcttc gctgattaat taccccagaa ataaggctaa
      181 aaaactaatc gcattattat cctatggttg ttaatttgat tcgttgattt gaaggtttgt
      241 ggggccaggt tactgccaat ttttcctctt cataaccata aaagctagta ttgtagaatc
      301 tttattgttc ggagcagtgc ggcgcgaggc acatctgcgt ttcaggaacg cgaccggtga
      361 agaccaggac gcacggagga gagtcttccg tcggagggct gtcgcccgct cggcggcttc
      421 taatccgtac ttcaatatag caatgagcag ttaagcgtat tactgaaagt tccaaagaga
      481 aggttttttt aggctaagat aatggggctc tttacatttc cacaacatat aagtaagatt
      541 agatatggat atgtatatgg tggtattgcc atgtaatatg attattaaac ttctttgcgt
      601 ccatccaaaa aaaaaagtaa gaatttttga aaattcaata taaggatcca aaatggcttc
      661 agaaaaagaa attaggagag agagattctt gaacgttttc cctaaattag tagaggaatt
      721 gaacgcatcg cttttggctt acggtatgcc taaggaagca tgtgactggt atgcccactc
      781 attgaactac aacactccag gcggtaagtt aaatagaggt ttgtccgttg tggacacgta
      841 tgctattctc tccaacaaga ccgttgaaca attggggcaa gaagaatacg aaaaggttgc
      901 tattctaggt tggtgcattg agttgttgca ggcttacttc ttggtcgccg atgatatgat
      961 ggacaagtcc attaccagaa gaggccaacc atgttggtac aaggttcctg aagttgggga
     1021 aattgccatc aatgacgcat tcatgttaga ggctgctatc tacaagcttt tgaaatctca
     1081 cttcagaaac gaaaaatact acatagatat caccgaattg ttccatgaag tcaccttcca
     1141 aaccgaattg ggccaattga tggacttaat cactgcacct gaagacaaag tcgacttgag
     1201 taagttctcc ctaaagaagc actccttcat agttactttc aagactgctt actattcttt
     1261 ctacttgcct gtcgcattgg ctatgtacgt tgccggtatc acagatgaaa aggatttgaa
     1321 acaagccaga gatgtcttga ttccattggg tgaatatttc caaattcaag atgactactt
     1381 agactgcttc ggtaccccag aacagatcgg taagatcggt acagatatcc aagataacaa
     1441 atgttcttgg gtaatcaaca aggcattaga acttgcttcc gcagaacaaa gaaagacttt
     1501 agacgaaaat tacggtaaga aggactcagt cgcagaagcc aaatgcaaaa agattttcaa
     1561 tgacttgaaa atcgaccagt tataccacga atatgaagag tctgttgcca aggatttgaa
     1621 ggccaagatc tcccaagtcg acgagtctcg tggcttcaaa gccgacgtct taactgcgtt
     1681 tttgaacaaa gtttacaaga gaagcaaata aagatctttt gccagcttac tatccttctt
     1741 gaaaatatgc actctatatc ttttagttct taattgcaac acatagattt gctgtataac
     1801 gaattttatg ctatttttta aatttggagt tcagtgataa aagtgtcaca gcgaatttcc
     1861 tcacatgtag ggaccgaatt gtttacaagt tctctgtacc accatggaga catcaaaaat
     1921 tgaaaatcta tggaaagata tggacggtag caacaagaat atagcacgag ccgcggagtt
     1981 catttcgtta cttttgatat cactcacaac tattgcgaag cgcttcagtg aaaaaatcat
     2041 aaggaaaagt tgtaaatatt attggtagta ttcgtttggt aaagtagagg gggtaatttt
     2101 tcccctttat tttgttcata cattcttaaa ttgctttgcc tctccttttg gaaagctata
     2161 cttcggagca ctgttgagcg aaggctcatt agatatattt tctgtcattt tccttaaccc
     2221 aaaaataagg gaaagggtcc aaaaagcgct cggacaactg ttgaccgtga tccgaaggac
     2281 tggctataca gtgttcacaa aatagccaag ctgaaaataa tgtgtagcta tgttcagtta
     2341 gtttggctag caaagatata aaagcaggtc ggaaatattt atgggcatta ttatgcagag
     2401 catcaacatg ataaaaaaaa acagttgaat attccctcaa aaagctcaaa atgggatcct
     2461 ctaaaggtga agaattattc actggtgttg tcccaatttt ggttgaatta gatggtgatg
     2521 ttaatggtca caaattttct gtctccggtg aaggtgaagg tgatgctact tacggtaaat
     2581 tgaccttaaa atttatttgt actactggta aattgccagt tccatggcca accttagtca
     2641 ctactttcgg ttatggtgtt caatgttttg cgagataccc agatcatatg aaacaacatg
     2701 actttttcaa gtctgccatg ccagaaggtt atgttcaaga aagaactatt tttttcaaag
     2761 atgacggtaa ctacaagacc agagctgaag tcaagtttga aggtgatacc ttagttaata
     2821 gaatcgaatt aaaaggtatt gattttaaag aagatggtaa cattttaggt cacaaattgg
     2881 aatacaacta taactctcac aatgtttaca tcatggctga caaacaaaag aatggtatca
     2941 aagttaactt caaaattaga cacaacattg aagatggttc tgttcaatta gctgaccatt
     3001 atcaacaaaa tactccaatt ggtgatggtc cagtcttgtt accagacaac cattacttat
     3061 ccactcaatc tgccttatcc aaagatccaa acgaaaagag agaccacatg gtcttgttag
     3121 aatttgttac tgctgctggt attacccatg gtatggatga attgtacaaa agatctgctg
     3181 gtggaggagg tactggtggt gctgagctca tggctaccgc agcaggtcct atcgcaacta
     3241 acaactcccc acaaaactcc aacgcttaca gaactccaat cgctccttcc gtaccaatta
     3301 ctcataaatg gtctatagct gaagatttga catgtatttc caatcctagt aagcacaata
     3361 accctcaaac tggttacaga tcattttctg acgaattata cgttaagtac gaagaaaagt
     3421 tggaagatgt tagaaaagca ttaagagaag ttgaagaaaa ccctttggaa ggtttagtta
     3481 tgatagacgc tttgcaaaga ttgggtatcg attaccattt cagaggtgaa attggtgcat
     3541 tcttgcaaaa gcaacaaatc atatcttcaa ctccagatgg ttaccctgaa catggtttgt
     3601 acgaagtttc aacattgttt agattcttaa gacaagaagg tcacaatgtt accgctgacg
     3661 tctttaataa cttcaaggat aaggaaggta gattcagatc agaattgtca acagatatta
     3721 gaggtttgat gtccttatac gaagcaagtc aattgagaat agaaggtgaa gacatcttag
     3781 atcaagctgc tgatttctcc agtcaattgt taggtagatg gacaaaagat cctaatcatc
     3841 acgaagccag attggtttct aacactttaa cacatccata ccacaagtca ttggctacct
     3901 ttatgggtca aaaattgtcc tacatgaact gcaagggtcc aaactgggac ggtgtcgata
     3961 atttgcaaga attagctaag atggatttga ctatcgtaca aagtatccat caaaaggaag
     4021 tattccaagt ttctcaatgg tggaaggata caggtttagc caatgaattg aaattggcta
     4081 gaaaccaacc attgaagtgg tatatgtggc ctatggccgc tttaaccgat ccaagattca
     4141 gtgaagaaag agttgaatta actaagccta tttcttttat atacatcata gatgacatct
     4201 tcgacgtcta tggtaccatt gaagaattga ccttgtttac tgatgcagtt aatagatggg
     4261 aattgtctgc cgtcgaacaa ttaccagact acatgaaagt atgtttcaag gctttgtacg
     4321 atgttaccaa cgaaatcgca tacaaaatct ataaaaagca tggtcaaaac cctattgatt
     4381 ccttgcaaaa gacttgggct agtttgtgca atgcattttt agttgaagca aagtggttcg
     4441 cctctggtca cttgccaaat gcagaagaat acttaaagaa cggtatcatc tcttcaggtg
     4501 ttcatgttgt cttggcccac atgtttttct tgttaggtga cggtattaca caagaatcag
     4561 ttgatttggt agatgactat ccaggtattt ccacaagtat cgcaaccatt ttgagattat
     4621 ctgatgactt gggttcagcc aaagatgaag accaagatgg ttatgatggt tcttacatcg
     4681 aatgttacat gaaggaacat aagggttcca gtgtcgattc agccagagaa gaagtaataa
     4741 gaatgatctc cgaagcatgg aaatgtttga ataaggaatg cttatcacca aacccttttt
     4801 ctgaatcatt cagaataggt tccttgaata tggctagaat gatccctatg atgtactctt
     4861 acgatgacaa ccataacttg ccaattttag aagaacacat gaaggcaatg atctataaca
     4921 catctttata actcgagata gattaattta aacagtatat gtacagtttt atatatatat
     4981 atatatatat acatatataa agaaacctgt gcgttttttg tattttcaaa tacatttagt
     5041 tttggcgcca cttctataaa aggtctcatg atattgttac tctagagatc ccaatacaac
     5101 agatcacgtg atctttttgt aagatgaagt tgaagtagtg ttgcaccgtg ccaatgcagg
     5161 tggctattag attaaatatg tgatttgttc tattaagttt cctgtataat taatggggag
     5221 cgctgattct cttttggtac gcttcccatc cagcatttct gtatctttca ccttcaacct
     5281 taggatctct acccttggcg aaaagtcctc tgccaacaat gatgatatct gatccaccac
     5341 ttacaacttc gtcgacggtt ctgtactgct gacccaatgc atcgcctttg tcgtctaaac
     5401 ctacacctgg ggtcatgatt agccaatcaa acccttcttc tcttcctccc atatcgttct
     5461 gagcaatgaa cccaataacg aaatctttat cactctttgc aatatcaacg gtacccttag
     5521 tatattcacc gtgtgctaga gaacccttgg aagacaattc agcaagcatc aataatcccc
     5581 ttggttcttt ggtgacctct tgcgcacctt gtttcaagcc agcaacaata ccagcaccag
     5641 taaccccgtg ggcgttggtg atatcagacc attctgcgat acggtaaacg cccgatgtat
     5701 attgtaattt gactgtgtta ccgatatcgg cgaattttct gtcctcaaat atcaagaact
     5761 tgtatttctc tgccaatgct ttcaatggaa cgacagtacc ctcataactg aaatcatcca
     5821 agatatcaac gtgtgttttc aaaaggcaaa tgtatggacc caacgtttca acaagtttca
     5881 atagctcatc agtcgaacga acgtcaagag aagcacacaa attggtcttc ttttcatcca
     5941 ttaaacgtaa aagtttcgat gcaaccggac ttgcatgagt ctcagctcta ctggtatatg
     6001 attttgtgga catgttttat cttcgtttcc tgcaggtttt tgttctgtgc agttgggtta
     6061 agaatactgg gcaatttcat gtttcttcaa cactacatat gcgtatatat accaatctaa
     6121 gtctgtgctc cttccttcgt tcttccttct gttcggagat taccgaatca aaaaaattta
     6181 aatgaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca
     6241 acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg
     6301 caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc tgatgcggta
     6361 ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc tcagtacaat
     6421 ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc
     6481 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     6541 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt
     6601 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     6661 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     6721 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     6781 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     6841 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     6901 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     6961 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     7021 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     7081 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     7141 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     7201 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     7261 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     7321 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     7381 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     7441 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     7501 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     7561 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     7621 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     7681 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     7741 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     7801 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     7861 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     7921 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     7981 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     8041 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     8101 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     8161 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     8221 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     8281 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     8341 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     8401 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     8461 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     8521 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     8581 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
     8641 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
     8701 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
     8761 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa
     8821 gcttgcatgc atttaaatca ttacgaccga gattcccggg taataactga tataattaaa
     8881 ttgaagctct aatttgtgag tttagtatac atgcatttac ttataataca gttttttagt
     8941 tttgctggcc gcatcttctc aaatatgctt cccagcctgc ttttctgtaa cgttcaccct
     9001 ctaccttagc atcccttccc tttgcaaata gtcctactag ttatagtttt ttctccttga
     9061 cgttaaa
//
