LOCUS       sequence-370744-        8244 bp    DNA     circular SYN 15-APR-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 8244)
  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 8244)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: pAF9D92
FEATURES             Location/Qualifiers
     source          1..8244
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             704..724
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3079..3098)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3080..3098)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3108..3125)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3108..3124)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3117..3139)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3265..3720
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3352..3371)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3562..3583
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3823..4497)
                     /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(4498..4778)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(4799..4818)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4918..4940
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4978..4996)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    5037..5540
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        5577..5681
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5682..5750,5751..6542)
                     /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(5900..5919)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6713..7301
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7202..7221
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7455..7472
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7589..7610
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7625..7630,7631..7648,7649..7655)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7663..7679
                     /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     7668..7690
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7687..7703
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7687..7703
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7722..7742
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7724..7742
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 gacaacaacc gccactccgc ggatcatgtc cagattggat aagtccaagg ttattaactc
       61 tgccttggaa ttgttgaacg aagttggtat tgaaggtttg accactagaa agttggctca
      121 aaaattgggt gttgaacaac ctaccttgta ctggcatgtt aagaacaaaa gagctttgtt
      181 ggacgctttg gccattgaaa tgttggatag acatcatacc catttctgtc cattggaagg
      241 tgaatcatgg caagatttct tgagaaacaa cgccaagtct ttcagatgtg ctttgttgtc
      301 tcatagagat ggtgctaaag ttcacttggg tactagacca actgaaaagc aatacgaaac
      361 tttggaaaac cagttggctt tcttgtgtca acaaggtttc tctttagaaa acgccttgta
      421 tgctttgtct gctgttggtc attttacctt gggttgtgtt ttggaagatc aagaacatca
      481 agtcgccaaa gaagaaaggg aaactccaac tactgattct atgccaccat tattgagaca
      541 agccattgag ttgtttgatc atcaaggtgc tgaaccagct tttttgtttg gtttggaatt
      601 gatcatctgc ggtttggaga aacagttgaa atgtgaatct ggctccggta ccggtggtgg
      661 tactccagct gctgcttcta ctttggaatc tagatgtacc ccacctaaga agaaaagaaa
      721 ggttgctagc actgccagcg tttcgaatac gcagaataag ctgaatgagc ttctcgatgc
      781 catcagacag gagtttctcc aagtctcaca agaggcaaat acctaccgtc ttcaaaacca
      841 aaaggattac gatttcaaaa tgaaccagca gctggctgag atgcagcaga taagaaacac
      901 cgtctacgaa ctggagctaa ctcacaggaa aatgaaggac gcgtacgaag aagagatcaa
      961 gcacttgaaa ctagggctgg agcaaagaga ccatcaaatt gcatctttga ccgtccagca
     1021 acagcagcaa cagcaacagc agcaacagca acagcagcaa caggtccagc agcatttaca
     1081 acagcaacaa cagcagctag ccgctgcatc tgcatctgtt ccagttgcgc aacaaccacc
     1141 ggctactact tcggccaccg ccactccagc agcaaacaca actactggtt cgccatcggc
     1201 cttcccagta caagctagcc gtcctaatct ggttggctca cagttgccta ccaccacttt
     1261 gcctgtggtg tcctcaaacg cccaacaaca actaccacaa cagcaactgc aacagcagca
     1321 acttcaacaa cagcaaccac ctccccaggt ttccgtggca ccattgagta acacagccat
     1381 caacggatct cctacttcta aagagaccac tactttaccc tctgtcaagg cacctgaatc
     1441 tacgttgaaa gaaactgaac cggaaaataa taatacctcg aagataaatg acaccggatc
     1501 cgccaccacg gccaccacta ccaccgcaac tgaaactgaa atcaaaccta aggaggaaga
     1561 cgccaccccg gctagtttgc accaggatca ctacttagtc ccttataatc aaagagcaaa
     1621 ccactctaaa cctatcccac ctttcctttt ggatctagat tcccagtctg ttcccgatgc
     1681 tctgaagaag caaacaaatg attattatat tttatacaac ccggcactac caagagaaat
     1741 tgacgttgag ttacacaaat ctttggatca tacttcagtt gtttgttgcg tgaagttcag
     1801 taacgatggt gaatacttag ccacaggctg caacaaaact actcaagtgt atcgcgtttc
     1861 agacggttct ctggtggccc gtttatctga cgattctgct gccaataacc atcgaaattc
     1921 gatcactgaa aataacacca ccacgtccac ggataacaat acaatgacaa ccactactac
     1981 caccacaatt actaccacag cgatgacttc ggcagcagaa ttggcaaaag atgtggaaaa
     2041 cctgaacact tcgtcttccc catcatccga cttgtatatc cgttcagtgt gtttttctcc
     2101 agatgggaaa tttttggcaa caggtgctga agacagactg attagaattt gggatattga
     2161 aaatagaaag attgttatga ttcttcaagg ccacgaacaa gatatttatt cattggacta
     2221 ctttccctca ggtgacaaat tagtctccgg ttctggtgac cgtaccgttc gtatttggga
     2281 cttacgtaca ggccagtgtt cattgacttt atccattgaa gatggtgtta ccaccgtcgc
     2341 tgtatcacca ggtgatggta aatacatcgc tgctggttct ctagatcgtg ctgtgagagt
     2401 ttgggattcc gagaccggat tcttggtgga aagactagat tcggaaaacg aatccggtac
     2461 aggccacaag gactctgttt atagcgttgt cttcacgaga gatggacaaa gcgttgtatc
     2521 cggctcatta gatagatctg ttaagctctg gaatttgcag aatgcaaaca acaagagcga
     2581 ttcgaaaact ccaaattccg gcacttgtga agttacgtat atcgggcata aagactttgt
     2641 attgtccgtg gccaccacac aaaatgatga gtacatcttg tccggttcca aagatcgtgg
     2701 tgtcctgttt tgggataaga aatccggcaa tccgttattg atgttgcaag gtcataggaa
     2761 ttcagttata tctgtggctg tggcaaacgg gtctccgctg ggtccagaat ataacgtttt
     2821 tgctactggt agcggtgatt gtaaagcaag gatttggaag tataaaaaaa tagcgccaaa
     2881 ttaacctgca ggatacccag ttgagaagac gagctatttt tctcatgttt cttttgttta
     2941 ttcacttttc tatacttttt cttttctatt gaaatatata ttatatctta tgtatgtttt
     3001 tttgtattat aaacaagcaa atttcgtctt gccatttacc ttctgatatt aatcacatgt
     3061 atatcgggcc ccaattcgcc ctatagtgag tcgtattacg cgcgctcact ggccgtcgtt
     3121 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
     3181 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     3241 ttgcgcagcc tgaatggcga atggacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     3301 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     3361 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     3421 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     3481 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     3541 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     3601 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     3661 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt
     3721 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tagatcggca
     3781 agtgcacaaa caatacttaa ataaatacta ctcagtaata acctatttct tagcattttt
     3841 gacgaaattt gctattttgt tagagtcttt tacaccattt gtctccacac ctccgcttac
     3901 atcaacacca ataacgccat ttaatctaag cgcatcacca acattttctg gcgtcagtcc
     3961 accagctaac ataaaatgta agctttcggg gctctcttgc cttccaaccc agtcagaaat
     4021 cgagttccaa tccaaaagtt cacctgtccc acctgcttct gaatcaaaca agggaataaa
     4081 cgaatgaggt ttctgtgaag ctgcactgag tagtatgttg cagtcttttg gaaatacgag
     4141 tcttttaata actggcaaac cgaggaactc ttggtattct tgccacgact catctccatg
     4201 cagttggacg atatcaatgc cgtaatcatt gaccagagcc aaaacatcct ccttaggttg
     4261 attacgaaac acgccaacca agtatttcgg agtgcctgaa ctatttttat atgcttttac
     4321 aagacttgaa attttccttg caataaccgg gtcaattgtt ctctttctat tgggcacaca
     4381 tataataccc agcaagtcag catcggaatc caaagcacat tctgcggcct ctgtgctctg
     4441 caagccgcaa actttcacca atggaccaga actacctgtg aaattaataa cagacatact
     4501 ccaagctgcc tttgtgtgct taatcacgta tactcacgtg ctcaatagtc accaatgccc
     4561 tccctcttgg ccctctcctt ttcttttttc gaccgaatta attcttaatc ggcaaaaaaa
     4621 gaaaagctcc ggatcaagat tgtacgtaag gtgacaagct atttttcaat aaagaatatc
     4681 ttccactact gccatctggc gtcataactg caaagtacac atatattacg atgctgttct
     4741 attaaatgct tcctatatta tatatatagt aatgtcgtga tctatggtgc actctcagta
     4801 caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg
     4861 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     4921 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc
     4981 tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct tagacggatc
     5041 gcttgcctgt aacttacacg cgcctcgtat cttttaatga tggaataatt tgggaattta
     5101 ctctgtgttt atttattttt atgttttgta tttggatttt agaaagtaaa taaagaaggt
     5161 agaagagtta cggaatgaag aaaaaaaaat aaacaaaggt ttaaaaaatt tcaacaaaaa
     5221 gcgtacttta catatatatt tattagacaa gaaaagcaga ttaaatagat atacattcga
     5281 ttaacgataa gtaaaatgta aaatcacagg attttcgtgt gtggtcttct acacagacaa
     5341 gatgaaacaa ttcggcatta atacctgaga gcaggaagag caagataaaa ggtagtattt
     5401 gttggcgatc cccctagagt cttttacatc ttcggaaaac aaaaactatt ttttctttaa
     5461 tttctttttt tactttctat ttttaattta tatatttata ttaaaaaatt taaattataa
     5521 ttatttttat agcacgtgat gaaaaggacc caggtggcac ttttcgggga aatgtgcgcg
     5581 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     5641 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     5701 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     5761 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     5821 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     5881 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     5941 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     6001 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     6061 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     6121 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     6181 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa
     6241 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     6301 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     6361 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     6421 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     6481 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     6541 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     6601 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     6661 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     6721 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     6781 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     6841 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
     6901 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     6961 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     7021 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     7081 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     7141 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     7201 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     7261 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     7321 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     7381 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     7441 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac
     7501 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact
     7561 ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc
     7621 aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat
     7681 ttcacacagg aaacagctat gaccatgatt acgccaagcg cgcaattaac cctcactaaa
     7741 gggaacaaaa gctggagctc tacaatgaaa aacaacacca agctgccaaa agctgtatat
     7801 taaagttaaa gctccctcat ttttagtaag gacgctttct tttcccacaa aagggacgag
     7861 gaaaaaaaga aaaacctgga aatatagtca cgtgacatga ataaacaaga cctagggcaa
     7921 tatggaaaaa gaagctagga taaaaataca tttatgaggg ttgtaaggca aagtggctca
     7981 gcattaagta gggctaaaaa gagtcgcaaa tatacatcgt accgagtgat gaacgtggtt
     8041 ttgaacacct tgttctcttt tgttctcgct ccctacattc actatatatt agaagaaatt
     8101 tcacctcaat ggacaactcg agaaatgaat acagaaatat gttttttagc gaaattttcc
     8161 tttcttcttg tcttcttgtt ttatttaaac ttccaaggct ttaactcagt gtcaaacata
     8221 acaacctcct cctcccccac ctac
//
