LOCUS       sequence-368101-        8969 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 8969)
  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 8969)
  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: pAF7E1
FEATURES             Location/Qualifiers
     source          1..8969
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1..504
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        541..645
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(646..714,715..1506)
                     /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(864..883)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1677..2265
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2166..2185
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2419..2436
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2553..2574
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2589..2594,2595..2612,2613..2619)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2627..2643
                     /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     2632..2654
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2651..2667
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2651..2667
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2686..2706
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2688..2706
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             3548..3568
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(7012..7031)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(7013..7031)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(7041..7058)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(7041..7057)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7050..7072)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      7198..7653
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7285..7304)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7495..7516
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(7756..8430)
                     /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(8431..8711)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(8732..8751)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8851..8873
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(8911..8929)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gatcgcttgc ctgtaactta cacgcgcctc gtatctttta atgatggaat aatttgggaa
       61 tttactctgt gtttatttat ttttatgttt tgtatttgga ttttagaaag taaataaaga
      121 aggtagaaga gttacggaat gaagaaaaaa aaataaacaa aggtttaaaa aatttcaaca
      181 aaaagcgtac tttacatata tatttattag acaagaaaag cagattaaat agatatacat
      241 tcgattaacg ataagtaaaa tgtaaaatca caggattttc gtgtgtggtc ttctacacag
      301 acaagatgaa acaattcggc attaatacct gagagcagga agagcaagat aaaaggtagt
      361 atttgttggc gatcccccta gagtctttta catcttcgga aaacaaaaac tattttttct
      421 ttaatttctt tttttacttt ctatttttaa tttatatatt tatattaaaa aatttaaatt
      481 ataattattt ttatagcacg tgatgaaaag gacccaggtg gcacttttcg gggaaatgtg
      541 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
      601 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
      661 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
      721 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
      781 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
      841 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
      901 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
      961 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     1021 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     1081 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     1141 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     1201 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     1261 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     1321 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     1381 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     1441 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     1501 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     1561 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     1621 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     1681 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     1741 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     1801 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     1861 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     1921 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     1981 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     2041 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     2101 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     2161 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     2221 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     2281 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     2341 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     2401 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     2461 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc
     2521 gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca
     2581 ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa
     2641 caatttcaca caggaaacag ctatgaccat gattacgcca agcgcgcaat taaccctcac
     2701 taaagggaac aaaagctgga gctctacaat gaaaacaaca ccaagctgcc aaaagctgta
     2761 tattaaagtt aaagctccct catttttagt aaggacgctt tcttttccca caaaagggac
     2821 gaggaaaaaa agaaaaacct ggaaatatag tcacgtgaca tgaataaaca agacctaggg
     2881 caatatggaa aaagaagcta ggataaaaat acatttatga gggttgtaag gcaaagtggc
     2941 tcagcattaa gtagggctaa aaagagtcgc aaatatacat cgtaccgagt gatgaacgtg
     3001 gttttgaaca ccttgttctc ttttgttctc gctccctaca ttcactatat attagaagaa
     3061 atttcacctc aatggacaac tcgagaaatg aatacagaaa tatgtttttt agcgaaattt
     3121 tcctttcttc ttgtcttctt gttttattta aacttccaag gctttaactc agtgtcaaac
     3181 ataacaacct cctcctcccc cacctacgac aacaaccgcc actggatcca tcatgggtac
     3241 tagaccatac gcttgtccag ttgaatcttg tgatagaaga ttctccagat ccgatgaatt
     3301 gaccagacat attagaatcc ataccggtca aaagccattc caatgtagaa tctgtatgag
     3361 gaacttctcc aggtctgatc atttgactac tcatatcaga acccacactg gtgaaaaacc
     3421 attcgcttgt gatatttgcg gtagaaagtt tgctagatcc gacgaaagaa aaagacatac
     3481 caaaattcat tctagaggtg gtggtactcc agctgctgct tctactttgg aagtcgactg
     3541 taccccacct aagaagaaaa gaaaggttgg tacctctgct gctcctgtcc aagacaaaga
     3601 cactctgtcc aatgccgagc gtgcgaagaa cgtcaacggg ttgcttcagg tgctcatgga
     3661 cattaacact ctgaacggag ggagctccga cactgctgat aagataagaa ttcatgccaa
     3721 aaacttcgag gcagctttgt tcgcaaagag ctcttcaaag aaagaataca tggacagcat
     3781 gaacgaaaaa gttgctgtca tgcgcaacac gtacaatacg aggaaaaacg ccgttactgc
     3841 tgctgccgct aataacaaca ttaaacccgt ggaacagcac catattaaca acttgaaaaa
     3901 ttctggcaac agcgccaata atatgaatgt caatatgaat ctaaacccac agatgttctt
     3961 gaatcagcag gctcaggcaa ggcaacaggt tgcgcaacaa ttaagaaatc aacaacaaca
     4021 acaacaacag cagcagcagc aacagaggcg tcaattgact cctcaacaac aacaattagt
     4081 gaaccagatg aaagtggcac ctattcccaa acaattactg caaagaattc ctaacattcc
     4141 acccaatatc aacacctggc agcaggtcac tgctttggct caacaaaagc tattgacacc
     4201 tcaggatatg gaagctgcga aggaagtcta caagattcac cagcagttgc tattcaaagc
     4261 aaggctacag caacaacaag cacaggctca agcacaagct aataacaaca acaacggcct
     4321 cccccaaaat ggtaatatta acaataacat aaatattcct caacagcagc aaatgcaacc
     4381 tcccaattca agtgcgaaca acaacccttt gcaacagcaa tcatcacaaa ataccgtacc
     4441 aaacgtcctc aaccaaatta accaaatctt ttctccagag gagcaacgca gcttattaca
     4501 agaagccatc gaaacctgca agaattttga aaaaacacaa ttgggtagta cgatgacgga
     4561 acctgtcaag caaagtttta ttaggaaata cattaaccaa aaggccctga gaaaaatcca
     4621 agctttgaga gatgttaaga acaacaataa cgctaacaac aacggctcga accttcagag
     4681 agcccaaaat gtccctatga atatcatcca acaacaacaa caacagaaca cgaacaataa
     4741 tgacaccatt gccacttctg ctacacctaa tgctgccgct ttctctcagc aacagaacgc
     4801 aagttctaaa ttatatcaga tgcaacaaca gcaacaagct caagctcaag ctcaagctca
     4861 agctcaggca caggctcaag cacaagctca agcacaggcg gcacaagcgg cgcaagcgca
     4921 agcacaagca caagcacaag cacaagcaca ggcacaggca caggcacaag cccaggcgca
     4981 ggcccaagcc caagcccaag cacaagcaca agcacacgct cagcaccagc cctcccaaca
     5041 accacaacaa gctcagcaac aacctaaccc actacatggg ttgacaccta ctgcaaagga
     5101 tgtcgaagta attaagcaat tgtccttgga tgcttctaag accaacctaa ggcttacgga
     5161 cgtaacaaat tctttatcca atgaagaaaa ggaaaaaatt aaaatgaagt taaagcaagg
     5221 tcaaaagctt tttgttcagg tgagtaattt cgccccacaa gtctacatca tcacaaagaa
     5281 tgaaaacttc ttgaaggaag tttttcagtt aagaatcttt gtaaaagaga tcctagaaaa
     5341 atgtgccgag ggtatatttg ttgttaaatt agacaccgtt gacaggttaa ttattaagta
     5401 tcaaaaatat tgggaaagta tgagaattca aattttaaga agacaagcta ttttaagaca
     5461 acaacagcaa atggcaaaca acaatgggaa cccaggcact acttctactg gaaacaataa
     5521 taatattgca actcagcaaa atatgcaaca gtcactacag caaatgcagc atttacagca
     5581 attgaaaatg cagcagcaac aacaacagca gcaacaacaa cagcagcaac aacaacagca
     5641 gcaacaacaa cagcaacagc acatatatcc ctcctcgact cctggtgtgg ctaattattc
     5701 ggcaatggct aatgcacccg gtaacaatat cccatatatg aaccataaaa atacctctag
     5761 catggatttt ttgaactcta tggaaaatac accaaaagtt cccgtatccg ctgcggcaac
     5821 cccatcactg aacaagacga tcaacggtaa ggtgaatggc aggacaaaat ctaatacgat
     5881 acctgttacc agcattccat caacaaataa gaaactatca atttcaaatg ccgctagtca
     5941 acaacccact cctaggtctg catcaaatac cgctaagtca accccaaata caaatccttc
     6001 tccactgaaa acccaaacta aaaatggcac accaaacccc aataatatga agactgtaca
     6061 gtctcctatg ggtgcacaac catcatataa tagtgccatt atagagaatg cattcagaaa
     6121 ggaagaactc ttgttaaaag atttggaaat caggaagttg gagatatctt ctcgttttaa
     6181 acatcgccaa gaaattttca aagattctcc tatggatttg tttatgagta cgctgggtga
     6241 ttgcttaggt atcaaagatg aagagatgct tacgtcatgc actatcccaa aggctgtggt
     6301 tgatcacatc aacggctctg gcaaaaggaa gcctacaaaa gcggctcaaa gggctcgcga
     6361 tcaagattcc attgacattt ccataaaaga caacaaattg gttatgaaga gtaaattcaa
     6421 taagagcaat aggtcgtatt cgatagcgtt gtccaatgta gctgctatat tcaagggtat
     6481 cggtggtaac tttaaagact tgtccacttt ggttcattca tcatcgccgt ccacatcttc
     6541 taatatggat gtcggcaacc caagaaaaag aaaagccagc gtattagaaa taagcccgca
     6601 agattcgata gcctcggtgc tatcaccaga ttcaaatata atgagtgatt ctaaaaaaat
     6661 taaagtagat tctcctgatg acccattcat gacaaaatca ggagccacaa ctagtgaaaa
     6721 acaagaagta acaaatgaag ctccattttt aacttctggg actagttcag aacaattcaa
     6781 tgtatgggat tggaataatt ggacaagtgc tacttaacct gcaggatacc cagttgagaa
     6841 gacgagctat ttttctcatg tttcttttgt ttattcactt ttctatactt tttcttttct
     6901 attgaaatat atattatatc ttatgtatgt ttttttgtat tataaacaag caaatttcgt
     6961 cttgccattt accttctgat attaatcaca tgtatatcgg gccccaattc gccctatagt
     7021 gagtcgtatt acgcgcgctc actggccgtc gttttacaac gtcgtgactg ggaaaaccct
     7081 ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc
     7141 gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggacg
     7201 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     7261 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     7321 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     7381 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     7441 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     7501 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     7561 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     7621 cgaattttaa caaaatatta acgcttacaa tttcctgatg cggtattttc tccttacgca
     7681 tctgtgcggt atttcacacc gcatagatcg gcaagtgcac aaacaatact taaataaata
     7741 ctactcagta ataacctatt tcttagcatt tttgacgaaa tttgctattt tgttagagtc
     7801 ttttacacca tttgtctcca cacctccgct tacatcaaca ccaataacgc catttaatct
     7861 aagcgcatca ccaacatttt ctggcgtcag tccaccagct aacataaaat gtaagctttc
     7921 ggggctctct tgccttccaa cccagtcaga aatcgagttc caatccaaaa gttcacctgt
     7981 cccacctgct tctgaatcaa acaagggaat aaacgaatga ggtttctgtg aagctgcact
     8041 gagtagtatg ttgcagtctt ttggaaatac gagtctttta ataactggca aaccgaggaa
     8101 ctcttggtat tcttgccacg actcatctcc atgcagttgg acgatatcaa tgccgtaatc
     8161 attgaccaga gccaaaacat cctccttagg ttgattacga aacacgccaa ccaagtattt
     8221 cggagtgcct gaactatttt tatatgcttt tacaagactt gaaattttcc ttgcaataac
     8281 cgggtcaatt gttctctttc tattgggcac acatataata cccagcaagt cagcatcgga
     8341 atccaaagca cattctgcgg cctctgtgct ctgcaagccg caaactttca ccaatggacc
     8401 agaactacct gtgaaattaa taacagacat actccaagct gcctttgtgt gcttaatcac
     8461 gtatactcac gtgctcaata gtcaccaatg ccctccctct tggccctctc cttttctttt
     8521 ttcgaccgaa ttaattctta atcggcaaaa aaagaaaagc tccggatcaa gattgtacgt
     8581 aaggtgacaa gctatttttc aataaagaat atcttccact actgccatct ggcgtcataa
     8641 ctgcaaagta cacatatatt acgatgctgt tctattaaat gcttcctata ttatatatat
     8701 agtaatgtcg tgatctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     8761 gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     8821 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac
     8881 cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta
     8941 atgtcatgat aataatggtt tcttagacg
//
