LOCUS       sequence-60746-e        6237 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 6237)
  AUTHORS   Nevozhay D, Adams RM, Murphy KF, Josic K, Balazsi G
  TITLE     Negative autoregulation linearizes the dose-response and suppresses 
            the heterogeneity of gene expression.
  JOURNAL   Proc Natl Acad Sci U S A. 2009 Mar 31;106(13):5123-8. doi: 
            10.1073/pnas.0809901106. Epub 2009 Mar 11.
  PUBMED    19279212
REFERENCE   2  (bases 1 to 6237)
  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"; doi: 
            "10.1073/pnas.0809901106"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2009-03-31- 31"; volume: "106"; issue: "13"; pages: "5123-8"
COMMENT     Sequence Label: pDN-G1Gh
FEATURES             Location/Qualifiers
     source          1..6237
                     /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        317..504
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             505..1167
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      complement(1427..1882)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1564..1585)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1776..1795
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(1965..2130)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     2051..2075
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    2625..2641
                     /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)."
     promoter        2772..3213
                     /label=TPGI promoter
                     /note="S. cerevisiae GAL1 promoter modified to contain two 
                     copies of the tet operator (Ellis et al., 2009)"
     protein_bind    2772..2889
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     protein_bind    3097..3115
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3118..3136
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     3184..3207
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             3237..3953
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      3986..4175
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4247..4264)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4418..5006)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4498..4517)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5177..5968,5969..6037))
                     /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     5800..5819
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6038..6142)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6210..6228
                     /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 accataaatt cccgttttaa gagcttggtg agcgctagga gtcactgcca ggtatcgttt
      241 gaacacggca ttagtcaggg aagtcataac acagtccttt cccgcaattt tctttttcta
      301 ttactcttgg cctcctctag tacactctat atttttttat gcctcggtaa tgattttcat
      361 tttttttttt ccacctagcg gatgactctt tttttttctt agcgattggc attatcacat
      421 aatgaattat acattatata aagtaatgtg atttcttcga agaatatact aaaaaatgag
      481 caggcaagat aaacgaaggc aaagatgaca gagcagaaag ccctagtaaa gcgtattaca
      541 aatgaaacca agattcagat tgcgatctct ttaaagggtg gtcccctagc gatagagcac
      601 tcgatcttcc cagaaaaaga ggcagaagca gtagcagaac aggccacaca atcgcaagtg
      661 attaacgtcc acacaggtat agggtttctg gaccatatga tacatgctct ggccaagcat
      721 tccggctggt cgctaatcgt tgagtgcatt ggtgacttac acatagacga ccatcacacc
      781 actgaagact gcgggattgc tctcggtcaa gcttttaaag aggccctagg ggccgtgcgt
      841 ggagtaaaaa ggtttggatc aggatttgcg cctttggatg aggcactttc cagagcggtg
      901 gtagatcttt cgaacaggcc gtacgcagtt gtcgaacttg gtttgcaaag ggagaaagta
      961 ggagatctct cttgcgagat gatcccgcat tttcttgaaa gctttgcaga ggctagcaga
     1021 attaccctcc acgttgattg tctgcgaggc aagaatgatc atcaccgtag tgagagtgcg
     1081 ttcaaggctc ttgcggttgc cataagagaa gccacctcgc ccaatggtac caacgatgtt
     1141 ccctccacca aaggtgttct tatgtagtga caccgattat ttaaagctgc agcatacgat
     1201 atatatacat gtgtatatat gtatacctat gaatgtcagt aagtatgtat acgaacagta
     1261 tgatactgaa gatgacaagg taatgcatca ttctatacgt gtcattctga acgaggcgcg
     1321 ctttcctttt ttctttttgc tttttctttt tttttctctt gaactcgacg gatctatgcg
     1381 gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggaaat tgtaagcgtt
     1441 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     1501 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     1561 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     1621 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     1681 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     1741 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     1801 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     1861 aatgcgccgc tacagggcgc gtcgcgccat tcgccattca ggctgcgcaa ctgttgggaa
     1921 gggcgatcgg tgcgggcctc ttcgctatta cgccagctga attggagcga cctcatgcta
     1981 tacctgagaa agcaacctga cctacaggaa agagttactc aagaataaga attttcgttt
     2041 taaaacctaa gagtcacttt aaaatttgta tacacttatt ttttttataa cttatttaat
     2101 aataaaaatc ataaatcata agaaattcgc ttatttagaa gtgtcaacaa cgtatctacc
     2161 aacgatttga cccttttcca tcttttcgta aatttctggc aaggtagaca agccgacaac
     2221 cttgattgga gacttgacca aacctctggc gaagaattgt taattaagag ctcttaagac
     2281 ccactttcac atttaagttg tttttctaat ccgcatatga tcaattcaag gccgaataag
     2341 aaggctggct ctgcaccttg gtgatcaaat aattcgatag cttgtcgtaa taatggcggc
     2401 atactatcag tagtaggtgt ttccctttct tctttagcga cttgatgctc ttgatcttcc
     2461 aatacgcaac ctaaagtaaa atgccccaca gcgctgagtg catataatgc attctcacta
     2521 gtgcggccgc cctttagtga gggttgaatt cgaattttca aaaattctta cttttttttt
     2581 ggatggacgc aaagaagttt aataatcata ttacatgtgt gaaattgtta tccgctcaca
     2641 attccacaca acatatctaa tcttacttat atgttgtgga aatgtaaaga gccccattat
     2701 cttagcctaa aaaaaccttc tctttggaac tttcagtaat acgcttaact gctcattgct
     2761 atattgaagt acggattaga agccgccgag cgggtgacag ccctccgaag gaagactctc
     2821 ctccgtgcgt cctcgtcttc accggtcgcg ttcctgaaac gcagatgtgc ctcgcgccgc
     2881 actgctccga acaataaaga ttctacaata ctagctttta tggttatgaa gaggaaaaat
     2941 tggcagtaac ctggccccac aaaccttcaa atgaacgaat caaattaaca accataggat
     3001 gataatgcga ttagtttttt agccttattt ctggggtaat taatcagcga agcgatgatt
     3061 tttgatctat taacagatat ataaatgcaa aaactgtccc tatcagtgat agagatctcc
     3121 ctatcagtga tagagatatt acttcttatt caaatgtaat aaaagtatca acaaaaaatt
     3181 gttaatatac ctctatactt taacgtcaag gagaaaaaac cccggatcct attaaaatgt
     3241 ctaaaggtga agaattattc actggtgttg tcccaatttt ggttgaatta gatggtgatg
     3301 ttaatggtca caaattttct gtctccggtg aaggtgaagg tgatgctact tacggtaaat
     3361 tgaccttaaa atttatttgt actactggta aattgccagt tccatggcca accttagtca
     3421 ctactttcgg ttatggtgtt caatgttttg cgagataccc agatcatatg aaacaacatg
     3481 actttttcaa gtctgccatg ccagaaggtt atgttcaaga aagaactatt tttttcaaag
     3541 atgacggtaa ctacaagacc agagctgaag tcaagtttga aggtgatacc ttagttaata
     3601 gaatcgaatt aaaaggtatt gattttaaag aagatggtaa cattttaggt cacaaattgg
     3661 aatacaacta taactctcac aatgtttaca tcatggctga caaacaaaag aatggtatca
     3721 aagttaactt caaaattaga cacaacattg aagatggttc tgttcaatta gctgaccatt
     3781 atcaacaaaa tactccaatt ggtgatggtc cagtcttgtt accagacaac cattacttat
     3841 ccactcaatc tgccttatcc aaagatccaa acgaaaagag agaccacatg gtcttgttag
     3901 aatttgttac tgctgctggt attacccatg gtatggatga attgtacaaa taactcgagt
     3961 aagcttggta ccgcggctag ctaagatccg ctctaaccga aaaggaagga gttagacaac
     4021 ctgaagtcta ggtccctatt tattttttta tagttatgtt agtattaaga acgttattta
     4081 tatttcaaat ttttcttttt tttctgtaca gacgcgtgta cgcatgtaac attatactga
     4141 aaaccttgct tgagaaggtt ttgggacgct cgaagatcca gctgcattaa tgaatcggcc
     4201 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     4261 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     4321 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     4381 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     4441 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     4501 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     4561 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     4621 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     4681 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     4741 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     4801 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga
     4861 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     4921 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     4981 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     5041 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5101 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5161 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5221 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5281 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     5341 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     5401 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     5461 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     5521 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     5581 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     5641 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     5701 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     5761 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     5821 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     5881 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     5941 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6001 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6061 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6121 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     6181 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtc
//
