LOCUS       sequence-60896-e        5320 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5320)
  AUTHORS   Nevozhay D, Adams RM, Van Itallie E, Bennett MR, Balazsi G
  TITLE     Mapping the environmental fitness landscape of a synthetic gene 
            circuit.
  JOURNAL   PLoS Comput Biol. 2012;8(4):e1002480. doi: 
            10.1371/journal.pcbi.1002480. Epub 2012 Apr 12.
  PUBMED    22511863
REFERENCE   2  (bases 1 to 5320)
  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: "PLoS Comput
            Biol."; date: "2012"; volume: "8(4):e1002480. doi"; pages: " 
            10.1371/journal.pcbi.1002480. Epub 2012 Apr 12"
COMMENT     Sequence Label: pDN-T2dMFot
FEATURES             Location/Qualifiers
     source          1..5320
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        191..472
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             473..1147
                     /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"
     rep_origin      complement(1250..1705)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1387..1408)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1599..1618
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      1802..1989
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1857..1881)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    2034..2052
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2076..2094
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             2290..2907
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESG"
     primer_bind     complement(2363..2382)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      3069..3258
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3330..3347)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3501..4089)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3581..3600)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4260..5051,5052..5120))
                     /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     4883..4902
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5121..5225)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5293..5311
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatagatc aacgacatta ctatatatat aatataggaa gcatttaata gaacagcatc
      241 gtaatatatg tgtactttgc agttatgacg ccagatggca gtagtggaag atattcttta
      301 ttgaaaaata gcttgtcacc ttacgtacaa tcttgatccg gagcttttct ttttttgccg
      361 attaagaatt aattcggtcg aaaaaagaaa aggagagggc caagagggag ggcattggtg
      421 actattgagc acgtgagtat acgtgattaa gcacacaaag gcagcttgga gtatgtctgt
      481 tattaatttc acaggtagtt ctggtccatt ggtgaaagtt tgcggcttgc agagcacaga
      541 ggccgcagaa tgtgctctag attccgatgc tgacttgctg ggtattatat gtgtgcccaa
      601 tagaaagaga acaattgacc cggttattgc aaggaaaatt tcaagtcttg taaaagcata
      661 taaaaatagt tcaggcactc cgaaatactt ggttggcgtg tttcgtaatc aacctaagga
      721 ggatgttttg gctctggtca atgattacgg cattgatatc gtccaactgc atggagatga
      781 gtcgtggcaa gaataccaag agttcctcgg tttgccagtt attaaaagac tcgtatttcc
      841 aaaagactgc aacatactac tcagtgcagc ttcacagaaa cctcattcgt ttattccctt
      901 gtttgattca gaagcaggtg ggacaggtga acttttggat tggaactcga tttctgactg
      961 ggttggaagg caagagagcc ccgaaagctt acattttatg ttagctggtg gactgacgcc
     1021 agaaaatgtt ggtgatgcgc ttagattaaa tggcgttatt ggtgttgatg taagcggagg
     1081 tgtggagaca aatggtgtaa aagactctaa caaaatagca aatttcgtca aaaatgctaa
     1141 gaaataggtt attactgagt agtatttatt taagtattgt ttgtgcactt gccgatctat
     1201 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcagga aattgtaagc
     1261 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     1321 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     1381 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     1441 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     1501 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     1561 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     1621 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     1681 cttaatgcgc cgctacaggg cgcgtccatt cgccattcag gctgcgcaac tgttgggaag
     1741 ggcgatcggt gcgggcctct tcgctattac gccagctgct taaggcgcca cttctaaata
     1801 agcgaatttc ttatgattta tgatttttat tattaaataa gttataaaaa aaataagtgt
     1861 atacaaattt taaagtgact cttaggtttt aaaacgaaaa ttcttattct tgagtaactc
     1921 tttcctgtag gtcaggttgc tttctcaggt atagtatgag gtcgctctta ttgaccacac
     1981 ctctaccggc agatccgcta gggataacag ggtaatatag atcaattcct cgatccctat
     2041 cagtgataga gagtcgacaa agtcgagttt ctcgatccct atcagtgata gagagtcgac
     2101 aaagtcgagt ttctcgatcg agaccactgc atgcatgtgc tctgtatgta tataaaactc
     2161 ttgttttctt cttttctcta aatattcttt ccttatacat taggtccttt gtagcataaa
     2221 ttactatact tctatagaca cgcaaacaca aatacacaca ctaaattacc ggatcaattc
     2281 gggggatcca tgtctagatt agataaaagt aaagtgatta acggcgcatt agagctgctt
     2341 aatggggtcg gaatcgaagg tttaacaacc cgtaaactcg cccagaagct aggtgtagag
     2401 cagcctacat tgtattggca tgtaaaaaat aagcgggctt tgctcgacgc cttacccatt
     2461 gagatgttag ataggcacca tactcacttt tgccctttag aaggggaaag ctggcaagat
     2521 tttttacgta ataacgctaa aagttttaga tgtgctttac taagtcatcg cgatggagca
     2581 aaagtacatt taggtacacg gcctacagaa aaacagtatg aaactctcga aaatcaatta
     2641 gcctttttat gccaacaagg tttttcacta gagaatgcat tatatgcact cagcgctgtg
     2701 gggcacttta ctttaggttg cgtattggaa gaacaagagc atcaagtcgc taaagaagaa
     2761 agggaaacac ctactactga tagtatgccg ccattattac gacaagctat cgaattattt
     2821 gatcgccaag gtgcagagcc agccttctta ttcggccttg aattgatcat atgcggatta
     2881 gaaaaacaac ttaaatgtga aagtgggggt ccagctgacg ccttggatga ctttgatttg
     2941 gatatgttgc ctgcagatgc tcttgacgat ttcgacctag acatgctacc cgccgacgca
     3001 ttagatgact ttgatttaga tatgttgcca ggttaactcg agtaagcttg gtaccgcggc
     3061 tagctaagat ccgctctaac cgaaaaggaa ggagttagac aacctgaagt ctaggtccct
     3121 atttattttt ttatagttat gttagtatta agaacgttat ttatatttca aatttttctt
     3181 ttttttctgt acagacgcgt gtacgcatgt aacattatac tgaaaacctt gcttgagaag
     3241 gttttgggac gctcgaagat ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     3301 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     3361 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     3421 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     3481 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     3541 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     3601 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     3661 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     3721 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     3781 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     3841 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     3901 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     3961 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     4021 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     4081 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     4141 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     4201 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     4261 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     4321 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     4381 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     4441 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     4501 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     4561 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     4621 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     4681 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     4741 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     4801 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     4861 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     4921 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     4981 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     5041 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     5101 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     5161 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     5221 ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca
     5281 ttaacctata aaaataggcg tatcacgagg ccctttcgtc
//
