LOCUS       sequence-369635-        8903 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 8903)
  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 8903)
  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: pAF9C3
FEATURES             Location/Qualifiers
     source          1..8903
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             735..755
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3770..3789)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3771..3789)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3799..3816)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3799..3815)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3808..3830)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3956..4411
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4043..4062)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4253..4274
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4514..5188)
                     /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(5189..5469)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(5490..5509)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5609..5631
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5669..5687)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    5728..6231
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        6268..6372
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6373..6441,6442..7233)
                     /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(6591..6610)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7404..7992
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7893..7912
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8146..8163
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8280..8301
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8316..8321,8322..8339,8340..8346)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8354..8370
                     /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     8359..8381
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8378..8394
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8378..8394
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     8413..8433
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        8415..8433
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 aaacataaca acctcctcct cccccaccta cgacaacaac cgccactccg cggatcatgt
       61 ccagattgga taagtccaag gttattaact ctgccttggc tttgggtaac gaagttggta
      121 ttgaagtttt gaccactaga aagttggctc aaaaattggg tgttgaacaa cctaccttgt
      181 actggcatgt taagaacaaa agagctttgt tggacgcttt ggccattgaa atgttggata
      241 gacatcatac ccatttctgt ccattggaag gtgaatcatg gcaagatttc ttgagaaaca
      301 acgccaagtc tttcagatgt gctttgttgt ctcatagaga tggtgctaaa gttcacttgg
      361 gtactagacc aactgaaaag caatacgaaa ctttggaaaa mcagttggct ttcttgtgtc
      421 aacaaggttt ctctttagaa aacgccttgt atgctttgtc tgctgttggt cattttacct
      481 tgggttgtgt tttggaagat caagaacatc aagtcgccaa agaagaaagg gaaactccaa
      541 ctactgattc tatgccacca ttattgagac aagccattga gttgtttgat catcaaggtg
      601 ctgaaccagc ttttttgttt ggtttggaat tgatcatctg cggtttggag aaacagttga
      661 aatgtgaatc tggctccggt accggtggtg gtactccagc tgctgcttct actttggaat
      721 ctagatgtac cccacctaag aagaaaagaa aggttgctag cagaacgtct ctcactgaac
      781 agtacgctcc ttctagtata caacatcagc agcagcaccc gcaaagtatt agtcctatag
      841 caaacacgca gtatggtgat atccctgtta gaccggaaat tgatttagat cctagcattg
      901 tgcctgtggt ccctgaaccc actgagccca tcgaaaacaa tatatcgctt aatgaggaag
      961 tcactttctt cgaaaaggcc aagaggtata tcggcaataa acatttatac actgagtttt
     1021 tgaaaatttt aaatttgtac tctcaagata tacttgatct tgacgattta gtggaaaagg
     1081 tagatttcta cttgggttcc aataaagaac tatttacgtg gttcaaaaac tttgttggct
     1141 accaagaaaa aaccaaatgt atcgagaata ttgttcatga aaaacataga ttggatttag
     1201 atttatgtga ggcatttggc ccaagttaca agaggctacc aaaaagtgac actttcatgc
     1261 catgctcagg tagggatgat atgtgttggg aagtcttgaa cgatgaatgg gttggacatc
     1321 ctgtatgggc ttccgaagat tcgggattta ttgctcatcg taaaaaccag tatgaggaaa
     1381 cactattcaa gatcgaagag gaaagacatg agtatgattt ttacattgaa tcaaatttaa
     1441 gaactattca atgtttggaa acaattgtaa ataagatcga gaacatgact gagaatgaaa
     1501 aagccaattt taaactgcct ccaggtcttg gtcatacttc aatgactatt tataaaaaag
     1561 tgataaggaa agtttatgat aaggaaaggg ggttcgagat tattgatgct ttgcatgagc
     1621 accctgcagt gacagcccca gtcgttctga aaagattaaa gcaaaaagac gaagaatgga
     1681 gaagagctca acgtgaatgg aataaagttt ggagggagtt agaacagaag gtttttttca
     1741 agtcattaga tcatttaggc ttaacattta aacaggctga caagaaatta ttaactacaa
     1801 agcagttgat atcagagatt agcagcatca aagttgatca aacaaacaaa aaaattcact
     1861 ggttaactcc taaaccaaag agccagttag attttgattt ccctgataaa aacattttct
     1921 atgatatctt gtgtttggct gacactttta taacccatac cacagcctat tctaatcccg
     1981 ataaagaaag attgaaagat ttactaaaat acttcatctc tttgtttttt tctatttctt
     2041 tggaaaaaat cgaagaatcg ttgtactccc ataagcaaaa cgtgtcagaa tctagcggat
     2101 ctgacgatgg cagttctatt gcatcaagga agaggcccta tcaacaagaa atgagtttac
     2161 ttgatatttt acacaggagc agatatcaaa agctaaagcg gtctaatgat gaagatggca
     2221 aagttcccca gctctctgaa ccacccgaag aagaacctaa taccattgag gaggaagagc
     2281 tcatcgatga agaagctaaa aatccgtggc taactgggaa tttagtggag gaagcaaact
     2341 ctcagggtat tatccaaaat cgcagtattt ttaatctatt cgctaatacg aatatttaca
     2401 tatttttccg tcattggaca acgatttatg agcggctttt ggaaattaag caaatgaatg
     2461 aaaaggtcac aaaggaaatc aacacaaggt cgacagttac ttttgccaaa gatctagatt
     2521 tattatcgag tcaactttcc gaaatgggac tagattttgt tggtgaagac gcgtacaaac
     2581 aggttttaag actgagtaga aggttgatta atgggggtct tgaacatcag tggtttgaag
     2641 agagtttgcg gcaagcttac aataacaaag cgtttaaact ctatacaatt gataaagtca
     2701 cccaatcgtt ggtaaagcat gctcatacct tgatgactga cgctaaaact gcagaaataa
     2761 tggctttgtt cgttaaagat agaaatgcct ccaccacgag tgcgaaggac caaattatct
     2821 atcgcttgca ggtgcgctca catatgtcca atacagaaaa tatgtttaga atagagtttg
     2881 ataaaagaac tctgcatgtt tccattcaat atattgcact tgatgatttg acactaaagg
     2941 aaccaaaggc agacgaagat aaatggaaat attatgtaac atcgtatgct ctgccacatc
     3001 caacagaagg cattccgcac gagaaactga agataccatt tttggaaagg ctcatcgaat
     3061 ttggacaaga tattgatgga acagaggtag atgaggagtt ttcccccgaa ggcatctctg
     3121 tatcgacatt gaaaattaag atccagccta taacctacca actgcatatt gaaaacgggt
     3181 cttacgatgt ttttacccgt aaggctacta acaaatatcc tactattgct aatgataata
     3241 ctcaaaaagg aatggtttct cagaagaagg agttgatatc aaaatttcta gactgtgcag
     3301 ttggtttgag aaataattta gatgaggctc aaaaattaag tatgcaaaaa aaatgggaga
     3361 acctaaaaga cagcatagta aagacgagtg ctggcaacca gggaatagag agtgaaaccg
     3421 aaaaaggtaa aattacgaaa caggagcagt cggataattt ggacagctct actgcaagtg
     3481 tactgcctgc atccattact actgtaccac aagatgataa tatagaaacg actgggaata
     3541 ctgaatcttc agacaagggg gctaagattc aataacctgc aggataccca gttgagaaga
     3601 cgagctattt ttctcatgtt tcttttgttt attcactttt ctatactttt tcttttctat
     3661 tgaaatatat attatatctt atgtatgttt ttttgtatta taaacaagca aatttcgtct
     3721 tgccatttac cttctgatat taatcacatg tatatcgggc cccaattcgc cctatagtga
     3781 gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     3841 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     3901 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggacgcg
     3961 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     4021 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     4081 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     4141 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     4201 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     4261 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     4321 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     4381 aattttaaca aaatattaac gcttacaatt tcctgatgcg gtattttctc cttacgcatc
     4441 tgtgcggtat ttcacaccgc atagatcggc aagtgcacaa acaatactta aataaatact
     4501 actcagtaat aacctatttc ttagcatttt tgacgaaatt tgctattttg ttagagtctt
     4561 ttacaccatt tgtctccaca cctccgctta catcaacacc aataacgcca tttaatctaa
     4621 gcgcatcacc aacattttct ggcgtcagtc caccagctaa cataaaatgt aagctttcgg
     4681 ggctctcttg ccttccaacc cagtcagaaa tcgagttcca atccaaaagt tcacctgtcc
     4741 cacctgcttc tgaatcaaac aagggaataa acgaatgagg tttctgtgaa gctgcactga
     4801 gtagtatgtt gcagtctttt ggaaatacga gtcttttaat aactggcaaa ccgaggaact
     4861 cttggtattc ttgccacgac tcatctccat gcagttggac gatatcaatg ccgtaatcat
     4921 tgaccagagc caaaacatcc tccttaggtt gattacgaaa cacgccaacc aagtatttcg
     4981 gagtgcctga actattttta tatgctttta caagacttga aattttcctt gcaataaccg
     5041 ggtcaattgt tctctttcta ttgggcacac atataatacc cagcaagtca gcatcggaat
     5101 ccaaagcaca ttctgcggcc tctgtgctct gcaagccgca aactttcacc aatggaccag
     5161 aactacctgt gaaattaata acagacatac tccaagctgc ctttgtgtgc ttaatcacgt
     5221 atactcacgt gctcaatagt caccaatgcc ctccctcttg gccctctcct tttctttttt
     5281 cgaccgaatt aattcttaat cggcaaaaaa agaaaagctc cggatcaaga ttgtacgtaa
     5341 ggtgacaagc tatttttcaa taaagaatat cttccactac tgccatctgg cgtcataact
     5401 gcaaagtaca catatattac gatgctgttc tattaaatgc ttcctatatt atatatatag
     5461 taatgtcgtg atctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     5521 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     5581 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     5641 tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     5701 gtcatgataa taatggtttc ttagacggat cgcttgcctg taacttacac gcgcctcgta
     5761 tcttttaatg atggaataat ttgggaattt actctgtgtt tatttatttt tatgttttgt
     5821 atttggattt tagaaagtaa ataaagaagg tagaagagtt acggaatgaa gaaaaaaaaa
     5881 taaacaaagg tttaaaaaat ttcaacaaaa agcgtacttt acatatatat ttattagaca
     5941 agaaaagcag attaaataga tatacattcg attaacgata agtaaaatgt aaaatcacag
     6001 gattttcgtg tgtggtcttc tacacagaca agatgaaaca attcggcatt aatacctgag
     6061 agcaggaaga gcaagataaa aggtagtatt tgttggcgat ccccctagag tcttttacat
     6121 cttcggaaaa caaaaactat tttttcttta atttcttttt ttactttcta tttttaattt
     6181 atatatttat attaaaaaat ttaaattata attattttta tagcacgtga tgaaaaggac
     6241 ccaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     6301 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     6361 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     6421 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     6481 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     6541 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     6601 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     6661 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     6721 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     6781 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     6841 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     6901 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     6961 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     7021 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     7081 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     7141 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     7201 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     7261 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     7321 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     7381 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     7441 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     7501 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     7561 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     7621 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     7681 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     7741 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     7801 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     7861 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     7921 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     7981 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     8041 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     8101 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     8161 gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat
     8221 taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt
     8281 aatgtgagtt agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt
     8341 atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat
     8401 tacgccaagc gcgcaattaa ccctcactaa agggaacaaa agctggagct ctacaatgaa
     8461 aacaacacca agctgccaaa agctgtatat taaagttaaa gctccctcat ttttagtaag
     8521 gacgctttct tttcccacaa aagggacgag gaaaaaaaga aaaacctgga aatatagtca
     8581 cgtgacatga ataaacaaga cctagggcaa tatggaaaaa gaagctagga taaaaataca
     8641 tttatgaggg ttgtaaggca aagtggctca gcattaagta gggctaaaaa gagtcgcaaa
     8701 tatacatcgt accgagtgat gaacgtggtt ttgaacacct tgttctcttt tgttctcgct
     8761 ccctacattc actatatatt agaagaaatt tcacctcaat ggacaactcg agaaatgaat
     8821 acagaaatat gttttttagc gaaattttcc tttcttcttg tcttcttgtt ttatttaaac
     8881 ttccaaggct ttaactcagt gtc
//
