LOCUS       sequence-240987-        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: pSV_229
FEATURES             Location/Qualifiers
     source          1..6269
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     32..51
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     285..302
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    419..440
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(455..460,461..478,479..485)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    493..509
                     /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     498..520
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     517..533
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     517..533
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        554..572
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     protein_bind    1540..1558
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1561..1579
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1581..1599
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1647..1665
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     complement(2389..2407)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2599..2618)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2600..2618)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2625..2642)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2625..2641)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2634..2656)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2782..3237
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2869..2888)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3079..3100
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3368..4171)
                     /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(4172..4387)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4411..4430)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4530..4552
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4590..4608)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4676..4780
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4781..4849,4850..5641)
                     /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(4999..5018)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(5812..6269,1..131)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
       61 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
      121 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
      181 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
      241 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
      301 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
      361 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
      421 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
      481 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
      541 acgccaagct cgaaattaac cctcactaaa gggaacaaaa gctggtacca tcatagagaa
      601 atcatatgaa aaatcaaata caactctggg cttggaggaa gtcattataa gatccggtta
      661 catgggtgtt tgtattgata acattccctc ccaatatagc tcttacaata atatgactac
      721 attctccaat tcaatttgct atgcaagaaa gaatttaagc tcggttccct tatacagaga
      781 cttggaaatt caactctcaa atattgcatc ttccagctcc aaaacccaat caagcgttgt
      841 cttgaacatt ttgaagttag cccaattaac ttcggttaat gttatacatc catatgtctt
      901 gatggcaact gtaatcctaa caatcctaat gtttctattt attctgtatg taactgtccc
      961 taagttacca ttcaaactag cggttaataa atttctactg ttgctaagtt caactatagt
     1021 tttgacatgg ggtattggtg cgatgtggac tcatgtaggg ataaatgcaa gttacagatt
     1081 ggttccatca tcaagtatga atataatcac tgtcaagaag ggcaagaagg cagcagtaat
     1141 ggcttggttt agctttgcat tcctcctttt agacagtgtg gttttatggt taatattttt
     1201 aagggatagg aaaagcttga aagatgaaat tgataatgtt ccatgtgccc aaaataggta
     1261 caataactac tcttcggatt catctacatt gcattccaaa gtttagctcg agggtcagat
     1321 tcattatcta caaagactgt aaaaagaaag tggaatattc attcatatca tattttttct
     1381 attaactgcc tggtttcttt taaatttttt attggttgtc gacttgaacg gagtgacaat
     1441 atatatatat atatatttaa taatgacatc attatctgta aatctgattc ttaatgctat
     1501 tctagttatg taagagtggt cccctaggat attgaagtat ccctatcagt gatagagaag
     1561 tccctatcag tgatagagaa tccctatcag tgatagagat gcgtcctcgt cttcaccggt
     1621 cgcgttcctg aaacgcagat gtgccttccc tatcagtgat agagaaacaa taaagattct
     1681 acaatactag cttttatggt tatgaagagg aaaaattggc agtaacctgg ccccacaaac
     1741 cttcaaatta acgaatcaaa ttaacaacca taggatgata atgcgattag ttttttagcc
     1801 ttatttctgg ggtaattaat cagcgaagcg atgatttttg atctattaac agatatataa
     1861 atggaaaagc tgcagatcta cccaggggtt gtttccatgg ctaaagaaat tggagattac
     1921 aaacgtacat tgtcatttaa gaaggttcat aatctgccaa ttacatttga tataacaaga
     1981 tgcttcccac attcaaacgt tacatgtcgt cctcagctca tcagattccc aaacatttta
     2041 agtcgctcat ctattcaact catgaagttg aggattgtac caaggttttg tcagtgaaaa
     2101 attatacgcc taaacaagac ttatctcaat caattgtgtt aaaaactttg gcctttccca
     2161 taaacccttc ggatatcaat cagttgcaag gagtataccc gtctcgtcca gaaaagacat
     2221 acgattactc cacagatgag ccggatccag ccgctatcgc cggtaatgag ggtgtctttg
     2281 aagttgtttc tttaccttcg ggaagttcca agggagattt gaaattgggt gaccgagtta
     2341 tcccattgga attctgtaca aatagggccg catcatgtaa ttagttatgt cacgcttaca
     2401 ttcacgccct ccccccacat ccgctctaac cgaaaaggaa ggagttagac aacctgaagt
     2461 ctaggtccct atttattttc ttatagttat gttagtatta agaacgttat ttatatttca
     2521 aatttttctt ttttttctgt acggacgcgt gtacgcatct agctagagcg gccgccaccg
     2581 cggtggagct ccaattcgcc ctatagtgag tcgtattaca attcactggc cgtcgtttta
     2641 caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc
     2701 cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg
     2761 cgcagcctga atggcgaatg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     2821 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     2881 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     2941 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     3001 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     3061 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     3121 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     3181 agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttcct
     3241 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatag ggtaataact
     3301 gatataatta aattgaagct ctaatttgtg agtttagtat acatgcattt acttataata
     3361 cagtttttta gttttgctgg ccgcatcttc tcaaatatgc ttcccagcct gcttttctgt
     3421 aacgttcacc ctctacctta gcatcccttc cctttgcaaa tagtcctctt ccaacaataa
     3481 taatgtcaga tcctgtagag accacatcat ccacggttct atactgttga cccaatgcgt
     3541 ctcccttgtc atctaaaccc acaccgggtg tcataatcaa ccaatcgtaa ccttcatctc
     3601 ttccacccat gtctctttga gcaataaagc cgataacaaa atctttgtcg ctcttcgcaa
     3661 tgtcaacagt acccttagta tattctccag tagataggga gcccttgcat gacaattctg
     3721 ctaacatcaa aaggcctcta ggttcctttg ttacttcttc tgccgcctgc ttcaaaccgc
     3781 taacaatacc tgggcccacc acaccgtgtg cattcgtaat gtctgcccat tctgctattc
     3841 tgtatacacc cgcagagtac tgcaatttga ctgtattacc aatgtcagca aattttctgt
     3901 cttcgaagag taaaaaattg tacttggcgg ataatgcctt tagcggctta actgtgccct
     3961 ccatggaaaa atcagtcaag atatccacat gtgtttttag taaacaaatt ttgggaccta
     4021 atgcttcaac taactccagt aattccttgg tggtacgaac atccaatgaa gcacacaagt
     4081 ttgtttgctt ttcgtgcatg atattaaata gcttggcagc aacaggacta ggatgagtag
     4141 cagcacgttc cttatatgta gctttcgaca tgatttatct tcgtttcctg caggtttttg
     4201 ttctgtgcag ttgggttaag aatactgggc aatttcatgt ttcttcaaca ctacatatgc
     4261 gtatatatac caatctaagt ctgtgctcct tccttcgttc ttccttctgt tcggagatta
     4321 ccgaatcaaa aaaatttcaa ggaaaccgaa atcaaaaaaa agaataaaaa aaaaatgatg
     4381 aattgaaaag gtggtatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     4441 ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     4501 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     4561 gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa
     4621 tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg
     4681 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     4741 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     4801 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     4861 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     4921 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     4981 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     5041 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     5101 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     5161 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     5221 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     5281 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     5341 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     5401 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     5461 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     5521 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     5581 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     5641 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     5701 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     5761 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     5821 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     5881 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     5941 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     6001 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     6061 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     6121 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     6181 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     6241 ggacaggtat ccggtaagcg gcagggtcg
//
