LOCUS       sequence-68820-e        5713 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  tetO-GFP reporter casette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5713)
  AUTHORS   Hsu C, Scherrer S, Buetti-Dinh A, Ratna P, Pizzolato J, Jaquet V, 
            Becskei A
  TITLE     Stochastic signalling rewires the interaction map of a multiple 
            feedback network during yeast evolution.
  JOURNAL   Nat Commun. 2012 Feb 21;3:682. doi: 10.1038/ncomms1687.
  PUBMED    22353713
REFERENCE   2  (bases 1 to 5713)
  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: "Nat 
            Commun."; date: "2012-02-21"; volume: "3:682. doi"; pages: " 
            10.1038/ncomms1687"
COMMENT     Sequence Label: pCH001
FEATURES             Location/Qualifiers
     source          1..5713
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        96..200
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(201..269,270..1061)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(419..438)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1232..1820
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1974..1991
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2107..2128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2143..2148,2149..2166,2167..2173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2181..2197
                     /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     2186..2208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2205..2221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2205..2221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2242..2260
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     terminator      2316..2503
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2371..2395)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    2548..2566
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             2765..3481
                     /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"
     primer_bind     complement(3524..3542)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(3729..3748)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3730..3748)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3755..3772)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3755..3771)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3764..3786)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3916..4371
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4003..4022)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4213..4234
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5146..5168
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        complement(5364..5584)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(5604..5623)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
      241 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
      301 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
      361 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
      421 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
      481 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
      541 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
      601 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttt cacaacatgg
      661 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
      721 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
      781 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
      841 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
      901 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
      961 cccgtatcgt agttatctac acgacgggca gtcaggcaac tatggatgaa cgaaatagac
     1021 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     1081 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     1141 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     1201 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     1261 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     1321 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     1381 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     1441 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     1501 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     1561 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     1621 agcattgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     1681 gcagggtcgg aacaggagag cgcacgaggg agcttccagg ggggaacgcc tggtatcttt
     1741 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     1801 gggggccgag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     1861 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     1921 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     1981 cagtgagcga ggaagcggaa gagcgcccaa acgcaaaccg cctctccccg cgcgttggcc
     2041 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
     2101 cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
     2161 ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga
     2221 ccatgattac gccaagctcg aaattaaccc tcactaaagg gaacaaaagc tggtaccggg
     2281 ccccccctcg agtaattcgc gccacttcta aataagcgaa tttcttatga tttatgattt
     2341 ttattattaa ataagttata aaaaaaataa gtgtatacaa attttaaagt gactcttagg
     2401 ttttaaaacg aaaattctta ttcttgagta actctttcct gtaggtcagg ttgctttctc
     2461 aggtatagta tgaggtcgct cttattgacc acacctctac cggcagatcc gctagggata
     2521 acagggtaat atagatcaat tcctcgatcc ctatcagtga tagagagtcg acaaagtcga
     2581 gtttctcgat cgagaccact gcatgcatgt gctctgtatg tatataaaac tcttgttttc
     2641 ttcttttctc taaatattct ttccttatac attaggtcct ttgtagcata aattactata
     2701 cttctataga cacgcaaaca caaatacaca cactaaatta ccggatcaat tcgggggatc
     2761 caaaatgtct aaaggtgaag aattattcac tggtgttgtc ccaattttgg ttgaattaga
     2821 tggtgatgtt aatggtcaca aattttctgt ctccggtgaa ggtgaaggtg atgctactta
     2881 cggtaaattg accttaaaat ttatttgtac tactggtaaa ttgccagttc catggccaac
     2941 cttagtcact actttcggtt atggtgttca atgttttgct agatacccag atcatatgaa
     3001 acaacatgac tttttcaagt ctgccatgcc agaaggttat gttcaagaaa gaactatttt
     3061 tttcaaagat gacggtaact acaagaccag agctgaagtc aagtttgaag gtgatacctt
     3121 agttaataga atcgaattaa aaggtattga ttttaaagaa gatggtaaca ttttaggtca
     3181 caaattggaa tacaactata actctcacaa tgtttacatc atggctgaca aacaaaagaa
     3241 tggtatcaaa gttaacttca aaattagaca caacattgaa gatggttctg ttcaattagc
     3301 tgaccattat caacaaaata ctccaattgg tgatggtcca gtcttgttac cagacaacca
     3361 ttacttatcc actcaatctg ccttatccaa agatccaaac gaaaagagag accacatggt
     3421 cttgttagaa tttgttactg ctgctggtat tacccatggt atggatgaat tgtacaaata
     3481 aaagaattct gtacaaatag ggccgcatca tgtaattagt tatgtcacgc ttacattcac
     3541 gccctccccc cacatccgct ctaaccgaaa aggaaggagt tagacaacct gaagtctagg
     3601 tccctattta tttttttata gttatgttag tattaagaac gttatttata tttcaaattt
     3661 ttcttttttt tctgtacgga cgcgtgtacg catctagagc ggccgcaccg cggtggagct
     3721 ccaattcgcc ctatagtgag tcgtattaca attcactggc cgtcgtttta caacgtcgtg
     3781 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cccttcgcca
     3841 gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga
     3901 atggcgaatg gcgcgacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     3961 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     4021 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     4081 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     4141 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     4201 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     4261 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     4321 tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt tcctgatgcg
     4381 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc agggtaataa ctgatataat
     4441 taaattgaag ctctaatttg tgagtttagt atacatgcat ttacttataa tacagttttt
     4501 tagttttgct ggccgcatct tctcaaatat gcttcccagc ctgcttttct gtaacgttca
     4561 ccctctacct tagcatccct tccctttgca aatagtcctc ttccaacaat aataatgtca
     4621 gatcctgtag agaccacatc atccacggtt ctatactgtt gacccaatgc gtctcccttg
     4681 tcatctaaac ccacaccggg tgtcataatc aaccaatcgt aaccttcatc tcttccaccc
     4741 atgtctcttt gagcaataaa gccgataaca aaatctttgt cgctcttcgc aatgtcaaca
     4801 gtacccttag tatattctcc agtagatagg gagcccttgc atgacaattc tgctaacatc
     4861 aaaaggcctc taggttcctt tgttacttct tctgccgcct gcttcaaacc gctaacaata
     4921 cctgggccca ccacaccgtg tgcattcgta atgtctgccc attctgctat tctgtataca
     4981 cccgcagagt actgcaattt gactgtatta ccaatgtcag caaattttct gtcttcgaag
     5041 agtaaaaaat tgtacttggc ggataatgcc tttagcggct taactgtgcc ctccatggaa
     5101 aaatcagtca agatatccac atgtgttttt agtaaatgac cgtctccggg agctgcatgt
     5161 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgacaaa ttttgggacc taatgcttca
     5221 actaactcca gtaattcctt ggtggtacga acatccaatg aagcacacaa gtttgtttgc
     5281 ttttcgtgca tgatattaaa tagcttggca gcaacaggac taggatgagt agcagcacgt
     5341 tccttatatg tagctttcga catgatttat cttcgtttcc tgcaggtttt tgttctgtgc
     5401 agttgggtta agaatactgg gcaatttcat gtttcttcaa cactacatat gcgtatatat
     5461 accaatctaa gtctgtgctc cttccttcgt tcttccttct gttcggagat taccgaatca
     5521 aaaaaatttc aaagaaaccg aaatcaaaaa aaagaataaa aaaaaaatga tgaattgaat
     5581 tgaaaagcgt ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc
     5641 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     5701 tacagacaag ctg
//
