LOCUS       sequence-68813-e        6927 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  rtTA under the control of GAL1 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6927)
  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 6927)
  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: pCH043
FEATURES             Location/Qualifiers
     source          1..6927
                     /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"
     promoter        join(2144..2149,2150..2167,2168..2174)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2182..2198
                     /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     2187..2209
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2206..2222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2206..2222
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2243..2261
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3230..3671
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    3230..3347
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     3642..3665
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             join(3689..4309,4310..4696)
                     /codon_start=1
                     /product="tetracycline-controlled transactivator, 
                     comprising a fusion of the tetracycline repressor TetR with
                     the C-terminal activation domain of herpes simplex virus 
                     VP16"
                     /label=tTA
                     /note="In the Tet-Off(R) system, tTA binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="MPRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTI
                     EGLLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHA
                     DALDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     primer_bind     complement(3762..3781)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(4737..4755)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(4943..4962)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4944..4962)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4969..4986)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4969..4985)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4978..5000)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5130..5585
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5217..5236)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5427..5448
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5714..6517)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(6518..6738)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(6758..6777)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6877..6899
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
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 tacgcaaacc gcctctcccc gcgcgttggc
     2041 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     2101 acgcaattaa tgtgagttac ctcactcatt aggcacccca ggctttacac tttatgcttc
     2161 cggctcctat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     2221 accatgatta cgccaagctc ggaattaacc ctcactaaag ggaacaaaag ctgggtacca
     2281 tcatagagaa atcatatgaa aaatcaaata caactctggg cttggaggaa gtcattataa
     2341 gatccggtta catgggtgtt tgtattgata acattccctc ccaatatagc tcttacaata
     2401 atatgactac attctccaat tcaatttgct atgcaagaaa gaatttaagc tcggttccct
     2461 tatacagaga cttggaaatt caactctcaa atattgcatc ttccagctcc aaaacccaat
     2521 caagcgttgt cttgaacatt ttgaagttag cccaattaac ttcggttaat gttatacatc
     2581 catatgtctt gatggcaact gtaatcctaa caatcctaat gtttctattt attctgtatg
     2641 taactgtccc taagttacca ttcaaactag cggttaataa atttctactg ttgctaagtt
     2701 caactatagt tttgacatgg ggtattggtg cgatgtggac tcatgtaggg ataaatgcaa
     2761 gttacagatt ggttccatca tcaagtatga atataatcac tgtcaagaag ggcaagaagg
     2821 cagcagtaat ggcttggttt agctttgcat tcctcctttt agacagtgtg gttttatggt
     2881 taatattttt aagggatagg aaaagcttga aagatgaaat tgataatgtt ccatgtgccc
     2941 aaaataggta caataactac tcttcggatt catctacatt gcattccaaa gtttagctcg
     3001 agggtcagat tcattatcta caaagactgt aaaaagaaag tggaatattc attcatatca
     3061 tattttttct attaactgcc tggtttcttt taaatttttt attggttgtc gacttgaacg
     3121 gagtgacaat atatatatat atatatttaa taatgacatc attatctgta aatctgattc
     3181 ttaatgctat tctagttatg taagagtggt ccgcatgcat attgaagtac ggattagaag
     3241 ccgccgagcg ggcgacagcc ctccgacgga agactctcct ccgtgcgtcc tcgtcttcac
     3301 cggtcgcgtt cctgaaacgc agatgtgcct cgcgccgcac tgctccgaac aataaagatt
     3361 ctacaatact agcttttatg gttatgaaga ggaaaaattg gcagtaacct ggccccacaa
     3421 accttcaaat taacgaatca aattaacaac cataggatga taatgcgatt agttttttag
     3481 ccttatttct ggggtaatta atcagcgaag cgatgatttt tgatctatta acagatatat
     3541 aaatggaaaa gctgcataac cactttaact aatactttca acattttcag tttgtattac
     3601 ttcttattca aatgtcataa aagtatcaac aaaaaattgt taatatacct ctatacttta
     3661 acgtcaagga gaaaaaacta taggatccat gcctagatta gataaaagta aagtgattaa
     3721 cagcgcatta gagctgctta atggggtcgg aatcgaaggt ttaacaaccc gtaaactcgc
     3781 ccagaagcta ggtgtagagc agcctacatt gtattggcat gtaaaaaata agcgggcttt
     3841 gctcgacgcc ttacccattg agatgttaga taggcaccat actcactttt gccctttaga
     3901 aggggaaagc tggcaagatt ttttacgtaa taacgctaaa agttttagat gtgctttact
     3961 aagtcatcgc gatggagcaa aagtacattt aggtacacgg cctacagaaa aacagtatga
     4021 aactctcgaa aatcaattag cctttttatg ccaacaaggt ttttcactag agaatgcatt
     4081 atatgcactc agcgctgtgg ggcactttac tttaggttgc gtattggaag aacaagagca
     4141 tcaagtcgct aaagaagaaa gggaaacacc tactactgat agtatgccgc cattattacg
     4201 acaagctatc gaattatttg atcgccaagg tgcagagcca gccttcttat tcggccttga
     4261 attgatcata tgcggattag aaaaacaact taaatgtgaa agtgggtccg cgtacagccg
     4321 cgcgcgtacg aaaaacaatt acgggtctac catcgagggc ctgctcgatc tcccggacga
     4381 cgacgccccc gaagaggcgg ggctggcggc tccgcgcctg tcctttctcc ccgcgggaca
     4441 cacgcgcaga ctgtcgacgg cccccccgac cgatgtcagc ctgggggacg agctccactt
     4501 agacggcgag gacgtggcga tggcgcatgc cgacgcgcta gacgatttcg atctggacat
     4561 gttgggggac ggggattccc cgggtccggg atttaccccc cacgactccg ccccctacgg
     4621 cgctctggat atggccgact tcgagtttga gcagatgttt accgatgccc ttggaattga
     4681 cgagtacggt gggtaggaat tctgtacaaa tagggccgca tcatgtaatt agttatgtca
     4741 cgcttacatt cacgccctcc ccccacatcc gctctaaccg aaaaggaagg agttagacaa
     4801 cctgaagtct aggtccctat ttattttttt atagttatgt tagtattaag aacgttattt
     4861 atatttcaaa tttttctttt ttttctgtac ggacgcgtgt acgcatctag agcggccgcc
     4921 accgcggtgg agctccaatt cgccctatag tgagtcgtat tacaattcac tggccgtcgt
     4981 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     5041 tccccccttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     5101 gttgcgcagc ctgaatggcg aatggcgcga cgcgccctgt agcggcgcat taagcgcggc
     5161 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     5221 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     5281 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     5341 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     5401 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     5461 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     5521 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac
     5581 aatttcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcagggta
     5641 ataactgata taattaaatt gaagctctaa tttgtgagtt tagtatacat gcatttactt
     5701 ataatacagt tttttagttt tgctggccgc atcttctcaa atatgcttcc cagcctgctt
     5761 ttctgtaacg ttcaccctct accttagcat cccttccctt tgcaaatagt cctcttccaa
     5821 caataataat gtcagatcct gtagagacca catcatccac ggttctatac tgttgaccca
     5881 atgcgtctcc cttgtcatct aaacccacac cgggtgtcat aatcaaccaa tcgtaacctt
     5941 catctcttcc acccatgtct ctttgagcaa taaagccgat aacaaaatct ttgtcgctct
     6001 tcgcaatgtc aacagtaccc ttagtatatt ctccagtaga tagggagccc ttgcatgaca
     6061 attctgctaa catcaaaagg cctctaggtt cctttgttac ttcttctgcc gcctgcttca
     6121 aaccgctaac aatacctggg cccaccacac cgtgtgcatt cgtaatgtct gcccattctg
     6181 ctattctgta tacacccgca gagtactgca atttgactgt attaccaatg tcagcaaatt
     6241 ttctgtcttc gaagagtaaa aaattgtact tggcggataa tgcctttagc ggcttaactg
     6301 tgccctccat ggaaaaatca gtcaagatat ccacatgtgt ttttagtaaa caaattttgg
     6361 gacctaatgc ttcaactaac tccagtaatt ccttggtggt acgaacatcc aatgaagcac
     6421 acaagtttgt ttgcttttcg tgcatgatat taaatagctt ggcagcaaca ggactaggat
     6481 gagtagcagc acgttcctta tatgtagctt tcgacatgat ttatcttcgt ttcctgcagg
     6541 tttttgttct gtgcagttgg gttaagaata ctgggcaatt tcatgtttct tcaacactac
     6601 atatgcgtat atataccaat ctaagtctgt gctccttcct tcgttcttcc ttctgttcgg
     6661 agattaccga atcaaaaaaa tttcaaagaa accgaaatca aaaaaaagaa taaaaaaaaa
     6721 atgatgaatt gaattgaaaa gcgtggtgca ctctcagtac aatctgctct gatgccgcat
     6781 agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     6841 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     6901 tttcaccgtc atcaccgaaa cgcgcga
//
