LOCUS       sequence-240996-        6269 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of the marked mRNA under the control of substitutional 
            promoters..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6269)
  AUTHORS   Baudrimont A, Voegeli S, Viloria EC, Stritt F, Lenon M, Wada T, 
            Jaquet V, Becskei A
  TITLE     Multiplexed gene control reveals rapid mRNA turnover.
  JOURNAL   Sci Adv. 2017 Jul 12;3(7):e1700006. doi: 10.1126/sciadv.1700006. 
            eCollection 2017 Jul.
  PUBMED    28706991
REFERENCE   2  (bases 1 to 6269)
  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: "Sci Adv."; 
            date: "2017-07-12"; volume: "3(7):e1700006. doi"; pages: " 
            10.1126/sciadv.1700006. eCollection 2017 Jul"
COMMENT     Sequence Label: pSM004
FEATURES             Location/Qualifiers
     source          1..6269
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    53..74
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(89..94,95..112,113..119)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    127..143
                     /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     132..154
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     151..167
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     151..167
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        188..206
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     protein_bind    1174..1192
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1195..1213
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1215..1233
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1281..1299
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     complement(2023..2041)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2233..2252)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2234..2252)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2259..2276)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2259..2275)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2268..2290)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2416..2871
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2503..2522)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2713..2734
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3002..3805)
                     /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(3806..4021)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4045..4064)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4164..4186
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4224..4242)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4310..4414
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4415..4483,4484..5275)
                     /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(4633..4652)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5446..6034
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5935..5954
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6188..6205
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
       61 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
      121 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
      181 agctcgaaat taaccctcac taaagggaac aaaagctggt accatcatag agaaatcata
      241 tgaaaaatca aatacaactc tgggcttgga ggaagtcatt ataagatccg gttacatggg
      301 tgtttgtatt gataacattc cctcccaata tagctcttac aataatatga ctacattctc
      361 caattcaatt tgctatgcaa gaaagaattt aagctcggtt cccttataca gagacttgga
      421 aattcaactc tcaaatattg catcttccag ctccaaaacc caatcaagcg ttgtcttgaa
      481 cattttgaag ttagcccaat taacttcggt taatgttata catccatatg tcttgatggc
      541 aactgtaatc ctaacaatcc taatgtttct atttattctg tatgtaactg tccctaagtt
      601 accattcaaa ctagcggtta ataaatttct actgttgcta agttcaacta tagttttgac
      661 atggggtatt ggtgcgatgt ggactcatgt agggataaat gcaagttaca gattggttcc
      721 atcatcaagt atgaatataa tcactgtcaa gaagggcaag aaggcagcag taatggcttg
      781 gtttagcttt gcattcctcc ttttagacag tgtggtttta tggttaatat ttttaaggga
      841 taggaaaagc ttgaaagatg aaattgataa tgttccatgt gcccaaaata ggtacaataa
      901 ctactcttcg gattcatcta cattgcattc caaagtttag ctcgagggtc agattcatta
      961 tctacaaaga ctgtaaaaag aaagtggaat attcattcat atcatatttt ttctattaac
     1021 tgcctggttt cttttaaatt ttttattggt tgtcgacttg aacggagtga caatatatat
     1081 atatatatat ttaataatga catcattatc tgtaaatctg attcttaatg ctattctagt
     1141 tatgtaagag tggtccccta ggatattgaa gtatccctat cagtgataga gaagtcccta
     1201 tcagtgatag agaatcccta tcagtgatag agatgcgtcc tcgtcttcac cggtcgcgtt
     1261 cctgaaacgc agatgtgcct tccctatcag tgatagagaa acaataaaga ttctacaata
     1321 ctagctttta tggttatgaa gaggaaaaat tggcagtaac ctggccccac aaaccttcaa
     1381 attaacgaat caaattaaca accataggat gataatgcga ttagtttttt agccttattt
     1441 ctggggtaat taatcagcga agcgatgatt tttgatctat taacagatat ataaatggaa
     1501 aagctgcaga tctaactgca taggcggcaa attcagccta aaagtttcca gaagcaggaa
     1561 ctcattccct attgattaat actcattaca aaaaccacaa tagagtagat aagatggcaa
     1621 aaacgttgaa ggatttacag ggttgggaga taataacaac tgatgaacaa ggtaacataa
     1681 tcgatggagg tcagaagaga ttacgccgaa gaggtgcaaa aactgaacat tacttaaaga
     1741 gaagttctga tggaattaaa ctaggtcgtg gtgatagtgt agtcatgcac aacgaagccg
     1801 ctgggactta ctccgtttat atgatccagg agttgagact taatacatta aataatgttg
     1861 tcgaactctg ggctctcaat tgcacctatt tacgatggtt tgaagtcaat cctttagctc
     1921 attataggca gtttaatcct gacgctaaca ttttgaatcg tcctttaaat tattacaata
     1981 aagaattctg tacaaatagg gccgcatcat gtaattagtt atgtcacgct tacattcacg
     2041 ccctcccccc acatccgctc taaccgaaaa ggaaggagtt agacaacctg aagtctaggt
     2101 ccctatttat tttcttatag ttatgttagt attaagaacg ttatttatat ttcaaatttt
     2161 tctttttttt ctgtacggac gcgtgtacgc atctagctag agcggccgcc accgcggtgg
     2221 agctccaatt cgccctatag tgagtcgtat tacaattcac tggccgtcgt tttacaacgt
     2281 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     2341 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     2401 ctgaatggcg aatggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     2461 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     2521 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     2581 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     2641 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     2701 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     2761 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     2821 tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt tcctgatgcg
     2881 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atagggtaat aactgatata
     2941 attaaattga agctctaatt tgtgagttta gtatacatgc atttacttat aatacagttt
     3001 tttagttttg ctggccgcat cttctcaaat atgcttccca gcctgctttt ctgtaacgtt
     3061 caccctctac cttagcatcc cttccctttg caaatagtcc tcttccaaca ataataatgt
     3121 cagatcctgt agagaccaca tcatccacgg ttctatactg ttgacccaat gcgtctccct
     3181 tgtcatctaa acccacaccg ggtgtcataa tcaaccaatc gtaaccttca tctcttccac
     3241 ccatgtctct ttgagcaata aagccgataa caaaatcttt gtcgctcttc gcaatgtcaa
     3301 cagtaccctt agtatattct ccagtagata gggagccctt gcatgacaat tctgctaaca
     3361 tcaaaaggcc tctaggttcc tttgttactt cttctgccgc ctgcttcaaa ccgctaacaa
     3421 tacctgggcc caccacaccg tgtgcattcg taatgtctgc ccattctgct attctgtata
     3481 cacccgcaga gtactgcaat ttgactgtat taccaatgtc agcaaatttt ctgtcttcga
     3541 agagtaaaaa attgtacttg gcggataatg cctttagcgg cttaactgtg ccctccatgg
     3601 aaaaatcagt caagatatcc acatgtgttt ttagtaaaca aattttggga cctaatgctt
     3661 caactaactc cagtaattcc ttggtggtac gaacatccaa tgaagcacac aagtttgttt
     3721 gcttttcgtg catgatatta aatagcttgg cagcaacagg actaggatga gtagcagcac
     3781 gttccttata tgtagctttc gacatgattt atcttcgttt cctgcaggtt tttgttctgt
     3841 gcagttgggt taagaatact gggcaatttc atgtttcttc aacactacat atgcgtatat
     3901 ataccaatct aagtctgtgc tccttccttc gttcttcctt ctgttcggag attaccgaat
     3961 caaaaaaatt tcaaggaaac cgaaatcaaa aaaaagaata aaaaaaaaat gatgaattga
     4021 aaaggtggta tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
     4081 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     4141 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     4201 accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     4261 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     4321 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     4381 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     4441 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     4501 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     4561 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     4621 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     4681 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     4741 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     4801 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     4861 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     4921 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     4981 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     5041 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     5101 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     5161 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     5221 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     5281 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     5341 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     5401 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     5461 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     5521 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     5581 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     5641 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     5701 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     5761 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     5821 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     5881 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     5941 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     6001 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     6061 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     6121 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     6181 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct
     6241 ccccgcgcgt tggccgattc attaatgca
//
