LOCUS       sequence-370739-        8703 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing artificial trans-activator in Saccharomyces cerevisiae.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8703)
  AUTHORS   Peng B, Bandari NC, Lu Z, Howard CB, Scott C, Trau M, Dumsday G, 
            Vickers CE
  TITLE     Engineering eukaryote-like regulatory circuits to expand artificial 
            control mechanisms for metabolic engineering in Saccharomyces 
            cerevisiae.
  JOURNAL   Commun Biol. 2022 Feb 16;5(1):135. doi: 10.1038/s42003-022-03070-z.
  PUBMED    35173283
REFERENCE   2  (bases 1 to 8703)
  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: "Commun 
            Biol."; date: "2022-02-16"; volume: "5(1):135. doi"; pages: " 
            10.1038/s42003-022-03070-z"
COMMENT     Sequence Label: pAF4E9D
FEATURES             Location/Qualifiers
     source          1..8703
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1513..1533
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3888..3907)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3889..3907)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3917..3934)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3917..3933)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3926..3948)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4074..4529
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4161..4180)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4371..4392
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4632..5306)
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     promoter        complement(5307..5587)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(5608..5627)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5727..5749
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5787..5805)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    5846..6349
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        6386..6490
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6491..6559,6560..7351)
                     /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(6709..6728)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7522..8110
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8011..8030
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8264..8281
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8398..8419
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8434..8439,8440..8457,8458..8464)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8472..8488
                     /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     8477..8499
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8496..8512
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8496..8512
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     8531..8551
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        8533..8551
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 acatgaataa acaagaccta gggcaatatg gaaaaagaag ctaggataaa aatacattta
       61 tgagggttgt aaggcaaagt ggctcagcat taagtagggc taaaaagagt cgcaaatata
      121 catcgtaccg agtgatgaac gtggttttga acaccttgtt ctcttttgtt ctcgctccct
      181 acattcacta tatattagaa gaaatttcac ctcaatggac aactcgagaa atgaatacag
      241 aaatatgttt tttagcgaaa ttttcctttc ttcttgtctt cttgttttat ttaaacttcc
      301 aaggctttaa ctcagtgtca aacataacaa cctcctcctc ccccacctac gacaacaacc
      361 gccactccgc ggatcatgaa gccagttacc ttgtacgatg ttgctgaata tgctggtgtt
      421 tcttaccaaa ccgtttccag agttgttaat caagcctctc atgtttctgc taagaccaga
      481 gaaaaagttg aagctgctat ggctgaattg aactacattc caaatagagt tgctcaacag
      541 ttggctggta aacagtcttt gttgatttgt gttgctactt cctctttggc tttacatgct
      601 ccatctcaaa tagttgctgc cattaagtca agagccgatc aattaggtgc ttccgttgtt
      661 gtttctatgg ttgaaagatc aggtgttgaa gcctgtaaag ctgctgttca taatttgttg
      721 gctcaaagag tttccggctt gatcattaac tatccattgg atgatcaaga tgccattgct
      781 gttgaagcag cttgtactaa tgttccagct ttgttcttgg atttctctga tcaaactcca
      841 atcaactcca tcatcttctc acatgaagat ggtactagat tgggtgttga acatttggtt
      901 gctttgggtc atcaacaaat tgctttgtta gctggtccat tgtcatctgt ttcagctaga
      961 ttgagattgg ctggttggca taagtacttg accagaaatc aaattcaacc tatcgctgaa
     1021 agagaaggcg attggtctgc tatgtctggt tttcaacaaa ccatgcagat gttgaacgaa
     1081 ggtatagttc caacagctat gttggttgct aacgatcaaa tggctttggg tgctatgaga
     1141 gctattactg aatctggttt gagagttggt gccgatattt ctgttgttgg ttacgatgat
     1201 accgaggatt cttcttgtta tattccacca ttgaccacca tcaagcaaga tttcagatta
     1261 ttgggtcaaa cctccgttga tagattgcta caattatctc aaggtcaagc cgttaagggt
     1321 aatcaattat tgccagtttc cttggtcaag agaaagacta ctttggctcc aaatactcaa
     1381 actgcttctc caagagcttt ggctgattct ttgatgcaat tagccagaca agtttccaga
     1441 ttggaatctg gtcaaggtac cggtggtggt actccagctg ctgcttctac tttggaatct
     1501 agatgtaccc cacctaagaa gaaaagaaag gttgctagca ctgccagcgt ttcgaatacg
     1561 cagaataagc tgaatgagct tctcgatgcc atcagacagg agtttctcca agtctcacaa
     1621 gaggcaaata cctaccgtct tcaaaaccaa aaggattacg atttcaaaat gaaccagcag
     1681 ctggctgaga tgcagcagat aagaaacacc gtctacgaac tggagctaac tcacaggaaa
     1741 atgaaggacg cgtacgaaga agagatcaag cacttgaaac tagggctgga gcaaagagac
     1801 catcaaattg catctttgac cgtccagcaa cagcagcaac agcaacagca gcaacagcaa
     1861 cagcagcaac aggtccagca gcatttacaa cagcaacaac agcagctagc cgctgcatct
     1921 gcatctgttc cagttgcgca acaaccaccg gctactactt cggccaccgc cactccagca
     1981 gcaaacacaa ctactggttc gccatcggcc ttcccagtac aagctagccg tcctaatctg
     2041 gttggctcac agttgcctac caccactttg cctgtggtgt cctcaaacgc ccaacaacaa
     2101 ctaccacaac agcaactgca acagcagcaa cttcaacaac agcaaccacc tccccaggtt
     2161 tccgtggcac cattgagtaa cacagccatc aacggatctc ctacttctaa agagaccact
     2221 actttaccct ctgtcaaggc acctgaatct acgttgaaag aaactgaacc ggaaaataat
     2281 aatacctcga agataaatga caccggatcc gccaccacgg ccaccactac caccgcaact
     2341 gaaactgaaa tcaaacctaa ggaggaagac gccaccccgg ctagtttgca ccaggatcac
     2401 tacttagtcc cttataatca aagagcaaac cactctaaac ctatcccacc tttccttttg
     2461 gatctagatt cccagtctgt tcccgatgct ctgaagaagc aaacaaatga ttattatatt
     2521 ttatacaacc cggcactacc aagagaaatt gacgttgagt tacacaaatc tttggatcat
     2581 acttcagttg tttgttgcgt gaagttcagt aacgatggtg aatacttagc cacaggctgc
     2641 aacaaaacta ctcaagtgta tcgcgtttca gacggttctc tggtggcccg tttatctgac
     2701 gattctgctg ccaataacca tcgaaattcg atcactgaaa ataacaccac cacgtccacg
     2761 gataacaata caatgacaac cactactacc accacaatta ctaccacagc gatgacttcg
     2821 gcagcagaat tggcaaaaga tgtggaaaac ctgaacactt cgtcttcccc atcatccgac
     2881 ttgtatatcc gttcagtgtg tttttctcca gatgggaaat ttttggcaac aggtgctgaa
     2941 gacagactga ttagaatttg ggatattgaa aatagaaaga ttgttatgat tcttcaaggc
     3001 cacgaacaag atatttattc attggactac tttccctcag gtgacaaatt agtctccggt
     3061 tctggtgacc gtaccgttcg tatttgggac ttacgtacag gccagtgttc attgacttta
     3121 tccattgaag atggtgttac caccgtcgct gtatcaccag gtgatggtaa atacatcgct
     3181 gctggttctc tagatcgtgc tgtgagagtt tgggattccg agaccggatt cttggtggaa
     3241 agactagatt cggaaaacga atccggtaca ggccacaagg actctgttta tagcgttgtc
     3301 ttcacgagag atggacaaag cgttgtatcc ggctcattag atagatctgt taagctctgg
     3361 aatttgcaga atgcaaacaa caagagcgat tcgaaaactc caaattccgg cacttgtgaa
     3421 gttacgtata tcgggcataa agactttgta ttgtccgtgg ccaccacaca aaatgatgag
     3481 tacatcttgt ccggttccaa agatcgtggt gtcctgtttt gggataagaa atccggcaat
     3541 ccgttattga tgttgcaagg tcataggaat tcagttatat ctgtggctgt ggcaaacggg
     3601 tctccgctgg gtccagaata taacgttttt gctactggta gcggtgattg taaagcaagg
     3661 atttggaagt ataaaaaaat agcgccaaat taacctgcag gatacccagt tgagaagacg
     3721 agctattttt ctcatgtttc ttttgtttat tcacttttct atactttttc ttttctattg
     3781 aaatatatat tatatcttat gtatgttttt ttgtattata aacaagcaaa tttcgtcttg
     3841 ccatttacct tctgatatta atcacatgta tatcgggccc caattcgccc tatagtgagt
     3901 cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg
     3961 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag
     4021 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggacgcgcc
     4081 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     4141 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     4201 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     4261 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     4321 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     4381 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     4441 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     4501 ttttaacaaa atattaacgc ttacaatttc ctgatgcggt attttctcct tacgcatctg
     4561 tgcggtattt cacaccgcat agatcggcaa gtgcacaaac aatacttaaa taaatactac
     4621 tcagtaataa cctatttctt agcatttttg acgaaatttg ctattttgtt agagtctttt
     4681 acaccatttg tctccacacc tccgcttaca tcaacaccaa taacgccatt taatctaagc
     4741 gcatcaccaa cattttctgg cgtcagtcca ccagctaaca taaaatgtaa gctttcgggg
     4801 ctctcttgcc ttccaaccca gtcagaaatc gagttccaat ccaaaagttc acctgtccca
     4861 cctgcttctg aatcaaacaa gggaataaac gaatgaggtt tctgtgaagc tgcactgagt
     4921 agtatgttgc agtcttttgg aaatacgagt cttttaataa ctggcaaacc gaggaactct
     4981 tggtattctt gccacgactc atctccatgc agttggacga tatcaatgcc gtaatcattg
     5041 accagagcca aaacatcctc cttaggttga ttacgaaaca cgccaaccaa gtatttcgga
     5101 gtgcctgaac tatttttata tgcttttaca agacttgaaa ttttccttgc aataaccggg
     5161 tcaattgttc tctttctatt gggcacacat ataataccca gcaagtcagc atcggaatcc
     5221 aaagcacatt ctgcggcctc tgtgctctgc aagccgcaaa ctttcaccaa tggaccagaa
     5281 ctacctgtga aattaataac agacatactc caagctgcct ttgtgtgctt aatcacgtat
     5341 actcacgtgc tcaatagtca ccaatgccct ccctcttggc cctctccttt tcttttttcg
     5401 accgaattaa ttcttaatcg gcaaaaaaag aaaagctccg gatcaagatt gtacgtaagg
     5461 tgacaagcta tttttcaata aagaatatct tccactactg ccatctggcg tcataactgc
     5521 aaagtacaca tatattacga tgctgttcta ttaaatgctt cctatattat atatatagta
     5581 atgtcgtgat ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     5641 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     5701 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     5761 atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     5821 catgataata atggtttctt agacggatcg cttgcctgta acttacacgc gcctcgtatc
     5881 ttttaatgat ggaataattt gggaatttac tctgtgttta tttattttta tgttttgtat
     5941 ttggatttta gaaagtaaat aaagaaggta gaagagttac ggaatgaaga aaaaaaaata
     6001 aacaaaggtt taaaaaattt caacaaaaag cgtactttac atatatattt attagacaag
     6061 aaaagcagat taaatagata tacattcgat taacgataag taaaatgtaa aatcacagga
     6121 ttttcgtgtg tggtcttcta cacagacaag atgaaacaat tcggcattaa tacctgagag
     6181 caggaagagc aagataaaag gtagtatttg ttggcgatcc ccctagagtc ttttacatct
     6241 tcggaaaaca aaaactattt tttctttaat ttcttttttt actttctatt tttaatttat
     6301 atatttatat taaaaaattt aaattataat tatttttata gcacgtgatg aaaaggaccc
     6361 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     6421 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     6481 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     6541 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     6601 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     6661 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     6721 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     6781 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     6841 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     6901 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     6961 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     7021 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     7081 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     7141 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     7201 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     7261 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     7321 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     7381 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     7441 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     7501 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     7561 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     7621 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     7681 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     7741 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     7801 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     7861 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     7921 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     7981 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     8041 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     8101 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     8161 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     8221 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     8281 ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta
     8341 atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa
     8401 tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat
     8461 gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta
     8521 cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctggagctct acaatgaaaa
     8581 acaacaccaa gctgccaaaa gctgtatatt aaagttaaag ctccctcatt tttagtaagg
     8641 acgctttctt ttcccacaaa agggacgagg aaaaaaagaa aaacctggaa atatagtcac
     8701 gtg
//
