LOCUS       sequence-370745-        7238 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 7238)
  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 7238)
  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: pAF9D93
FEATURES             Location/Qualifiers
     source          1..7238
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             671..691
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(2041..2060)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2042..2060)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2070..2087)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2070..2086)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2079..2101)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2227..2682
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2314..2333)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2524..2545
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2785..3459)
                     /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(3460..3740)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(3761..3780)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3880..3902
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3940..3958)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    3999..4502
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        4539..4643
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4644..4712,4713..5504)
                     /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(4862..4881)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5675..6263
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6164..6183
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6417..6434
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6551..6572
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6587..6592,6593..6610,6611..6617)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6625..6641
                     /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     6630..6652
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6649..6665
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6649..6665
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6684..6704
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6686..6704
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 attggataag tccaaggtta ttaactctgc cttggaattg ttgaacgaag ttggtattga
       61 aggtttgacc actagaaagt tggctcaaaa attgggtgtt gaacaaccta ccttgtactg
      121 gcatgttaag aacaaaagag ctttgttgga cgctttggcc attgaaatgt tggatagaca
      181 tcatacccat ttctgtccat tggaaggtga atcatggcaa gatttcttga gaaacaacgc
      241 caagtctttc agatgtgctt tgttgtctca tagagatggt gctaaagttc acttgggtac
      301 tagaccaact gaaaagcaat acgaaacttt ggaaaaccag ttggctttct tgtgtcaaca
      361 aggtttctct ttagaaaacg ccttgtatgc tttgtctgct gttggtcatt ttaccttggg
      421 ttgtgttttg gaagatcaag aacatcaagt cgccaaagaa gaaagggaaa ctccaactac
      481 tgattctatg ccaccattat tgagacaagc cattgagttg tttgatcatc aaggtgctga
      541 accagctttt ttgtttggtt tggaattgat catctgcggt ttggagaaac agttgaaatg
      601 tgaatctggc tccggtaccg gtggtggtac tccagctgct gcttctactt tggaatctag
      661 atgtacccca cctaagaaga aaagaaaggt tggtacctta aatacggcaa atttttcacc
      721 gccattacca cagcaacacc tatcgccttt aattcctatt gctattgctc cgaaagaaaa
      781 ttcaagtcga tcttctacaa gaaaaggtag aaaaaccaaa ttcgaaatcg gcgaaagtgg
      841 tgggaatgac ccatatatgg tttcttctcc caaaacgatg gctaagattc ccgtctcggt
      901 gaagcctcca ccttctttag cactgaataa tatgaactac caaacttcat ccgcttccac
      961 tgctttgtct tcgttgagca atagccatag tggcagtaga ctgaaactga acgcgttatc
     1021 gtccctacaa atgatgacgc ccattgctag cagtgcgcca aggactgttt tcatagacgg
     1081 tcctgaacag aaacaactac aacaacaaca aaattctctt tcaccacgtt attccaacac
     1141 tgttatatta ccaaggccgc gatctttaac ggattttcaa ggattgaaca atgcaaatcc
     1201 aaacaacaat ggaagtctca gagcacaaac tcagagttcc gtacagttga agagaccaag
     1261 ttcagtttta agtttgaacg acttgttggt tggccaaaga aataccaacg aatctgactc
     1321 tgattttact actggtggtg aggatgaaga agacggacta aaggacccgt ctaactctag
     1381 tatcgataac cttgagcaag actatttgca agagcaatca agaaagaaat ctaagacttc
     1441 cacgcccacg acaatgctaa gtagatccac tagtggtacg aatttgcaca ctttggggta
     1501 tgtaatgaac caaaatcact tgcatttctc ctcatcatct cctgatttcc aaaaggagtt
     1561 gaacaacaga ttactgaacg ttcaacaaca gcagcaagag caacataccc tactgcaatc
     1621 acaaaatacg tcaaaccaaa gtcaaaatca aaatcaaaat caaatgatgg cttccagtag
     1681 ttcgttaagt acaaccccgt tattattgtc accaagggtg aatatgatta atactgctat
     1741 atccacccaa caaaccccca tttctcagtc ggattcacaa gttcaagaac tggaaacatt
     1801 accacccata agaagtttac cgttgccctt cccacacatg gactaacctg caggataccc
     1861 agttgagaag acgagctatt tttctcatgt ttcttttgtt tattcacttt tctatacttt
     1921 ttcttttcta ttgaaatata tattatatct tatgtatgtt tttttgtatt ataaacaagc
     1981 aaatttcgtc ttgccattta ccttctgata ttaatcacat gtatatcggg ccccaattcg
     2041 ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg ttttacaacg tcgtgactgg
     2101 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     2161 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     2221 gaatggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     2281 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     2341 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     2401 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     2461 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     2521 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     2581 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     2641 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttcctgatgc ggtattttct
     2701 ccttacgcat ctgtgcggta tttcacaccg catagatcgg caagtgcaca aacaatactt
     2761 aaataaatac tactcagtaa taacctattt cttagcattt ttgacgaaat ttgctatttt
     2821 gttagagtct tttacaccat ttgtctccac acctccgctt acatcaacac caataacgcc
     2881 atttaatcta agcgcatcac caacattttc tggcgtcagt ccaccagcta acataaaatg
     2941 taagctttcg gggctctctt gccttccaac ccagtcagaa atcgagttcc aatccaaaag
     3001 ttcacctgtc ccacctgctt ctgaatcaaa caagggaata aacgaatgag gtttctgtga
     3061 agctgcactg agtagtatgt tgcagtcttt tggaaatacg agtcttttaa taactggcaa
     3121 accgaggaac tcttggtatt cttgccacga ctcatctcca tgcagttgga cgatatcaat
     3181 gccgtaatca ttgaccagag ccaaaacatc ctccttaggt tgattacgaa acacgccaac
     3241 caagtatttc ggagtgcctg aactattttt atatgctttt acaagacttg aaattttcct
     3301 tgcaataacc gggtcaattg ttctctttct attgggcaca catataatac ccagcaagtc
     3361 agcatcggaa tccaaagcac attctgcggc ctctgtgctc tgcaagccgc aaactttcac
     3421 caatggacca gaactacctg tgaaattaat aacagacata ctccaagctg cctttgtgtg
     3481 cttaatcacg tatactcacg tgctcaatag tcaccaatgc cctccctctt ggccctctcc
     3541 ttttcttttt tcgaccgaat taattcttaa tcggcaaaaa aagaaaagct ccggatcaag
     3601 attgtacgta aggtgacaag ctatttttca ataaagaata tcttccacta ctgccatctg
     3661 gcgtcataac tgcaaagtac acatatatta cgatgctgtt ctattaaatg cttcctatat
     3721 tatatatata gtaatgtcgt gatctatggt gcactctcag tacaatctgc tctgatgccg
     3781 catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
     3841 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     3901 ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt
     3961 tataggttaa tgtcatgata ataatggttt cttagacgga tcgcttgcct gtaacttaca
     4021 cgcgcctcgt atcttttaat gatggaataa tttgggaatt tactctgtgt ttatttattt
     4081 ttatgttttg tatttggatt ttagaaagta aataaagaag gtagaagagt tacggaatga
     4141 agaaaaaaaa ataaacaaag gtttaaaaaa tttcaacaaa aagcgtactt tacatatata
     4201 tttattagac aagaaaagca gattaaatag atatacattc gattaacgat aagtaaaatg
     4261 taaaatcaca ggattttcgt gtgtggtctt ctacacagac aagatgaaac aattcggcat
     4321 taatacctga gagcaggaag agcaagataa aaggtagtat ttgttggcga tccccctaga
     4381 gtcttttaca tcttcggaaa acaaaaacta ttttttcttt aatttctttt tttactttct
     4441 atttttaatt tatatattta tattaaaaaa tttaaattat aattattttt atagcacgtg
     4501 atgaaaagga cccaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     4561 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     4621 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     4681 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     4741 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     4801 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     4861 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     4921 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     4981 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     5041 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     5101 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     5161 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     5221 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     5281 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     5341 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     5401 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     5461 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     5521 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     5581 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     5641 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     5701 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     5761 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     5821 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     5881 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     5941 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     6001 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     6061 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     6121 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     6181 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     6241 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     6301 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     6361 gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg
     6421 agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt
     6481 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc
     6541 gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc
     6601 ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct
     6661 atgaccatga ttacgccaag cgcgcaatta accctcacta aagggaacaa aagctggagc
     6721 tctacaatga aaacaacacc aagctgccaa aagctgtata ttaaagttaa agctccctca
     6781 tttttagtaa ggacgctttc ttttcccaca aaagggacga ggaaaaaaag aaaaacctgg
     6841 aaatatagtc acgtgacatg aataaacaag acctagggca atatggaaaa agaagctagg
     6901 ataaaaatac atttatgagg gttgtaaggc aaagtggctc agcattaagt agggctaaaa
     6961 agagtcgcaa atatacatcg taccgagtga tgaacgtggt tttgaacacc ttgttctctt
     7021 ttgttctcgc tccctacatt cactatatat tagaagaaat ttcacctcaa tggacaactc
     7081 gagaaatgaa tacagaaata tgttttttag cgaaattttc ctttcttctt gtcttcttgt
     7141 tttatttaaa cttccaaggc tttaactcag tgtcaaacat aacaacctcc tcctccccca
     7201 cctacgacaa caaccgccac tccgcggatc atgtccag
//
