LOCUS       sequence-210283-        2893 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PTac-YFP Reporter.  IPTG sensor for use in Marionette strain..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2893)
  AUTHORS   Meyer AJ, Segall-Shapiro TH, Glassey E, Zhang J, Voigt CA
  TITLE     Escherichia coli ""Marionette"" strains with 12 highly optimized 
            small-molecule sensors.
  JOURNAL   Nat Chem Biol. 2018 Nov 26. pii: 10.1038/s41589-018-0168-3. doi: 
            10.1038/s41589-018-0168-3.
  PUBMED    30478458
REFERENCE   2  (bases 1 to 2893)
  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 Chem 
            Biol. 2018 Nov 26. pii: 10.1038/s41589-018-0168-3. doi: 
            10.1038/s41589-018-0168-3."
COMMENT     Sequence Label: pAJM.715
FEATURES             Location/Qualifiers
     source          1..2893
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        1..50
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(100..102,103..105,106..819)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(145..166)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(406..425)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     753..774
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     rep_origin      complement(1214..1759)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(1294..1313)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1848..2663)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1848..2663)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2571..2590
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2655..2674
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     promoter        join(2838..2843,2844..2859,2860..2866)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2872..2891
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
ORIGIN
        1 ctgtcaccgg atgtgctttc cggtctgatg agtccgtgag gacgaaacag cctctacaaa
       61 taattttgtt taatactaga gaaagagggg aaatactaga tggtgagcaa gggcgaggag
      121 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
      181 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
      241 atctgcacca caggcaagct gcccgtgccc tggcccaccc tcgtgaccac cttcggctac
      301 ggcctgcaat gcttcgcccg ctaccccgac cacatgaagc tgcacgactt cttcaagtcc
      361 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
      421 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
      481 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
      541 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
      601 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
      661 ccaatcggcg acggccccgt gctgctgccc gacaaccact accttagcta ccagtccgcc
      721 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
      781 gccgggatca ctctcggcat ggacgagctg tacaagtaac tcggtaccaa attccagaaa
      841 agaggcctcc cgaaaggggg gccttttttc gttttggtcc aatggcggcg cgccatcgaa
      901 tgaggtgctt cctcgctcac tgactcgctg cacgaggcag acctcagcgc tagcggagtg
      961 tatactggct tactatgttg gcactgatga gggtgtcagt gaagtgcttc atgtggcagg
     1021 agaaaaaagg ctgcaccggt gcgtcagcag aatatgtgat acaggatata ttccgcttcc
     1081 tcgctcactg actcgctacg ctcggtcgtt cgactgcggc gagcggaaat ggcttacgaa
     1141 cggggcggag atttcctgga agatgccagg aagatactta acagggaagt gagagggccg
     1201 cggcaaagcc gtttttccat aggctccgcc cccctgacaa gcatcacgaa atctgacgct
     1261 caaatcagtg gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggcg
     1321 gctccctcgt gcgctctcct gttcctgcct ttcggtttac cggtgtcatt ccgctgttat
     1381 ggccgcgttt gtctcattcc acgcctgaca ctcagttccg ggtaggcagt tcgctccaag
     1441 ctggactgta tgcacgaacc ccccgttcag tccgaccgct gcgccttatc cggtaactat
     1501 cgtcttgagt ccaacccgga aagacatgca aaagcaccac tggcagcagc cactggtaat
     1561 tgatttagag gagttagtct tgaagtcatg cgccggttaa ggctaaactg aaaggacaag
     1621 ttttggtgac tgcgctcctc caagccagtt acctcggttc aaagagttgg tagctcagag
     1681 aaccttcgaa aaaccgccct gcaaggcggt tttttcgttt tcagagcaag agattacgcg
     1741 cagaccaaaa cgatctcaag aagatcatct tattaagggg tctgacgctc agtggaacga
     1801 aaaatcaatc taaagtatat atgagtaaac ttggtctgac agttacctta gaaaaactca
     1861 tcgagcatca aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga
     1921 aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga
     1981 tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc
     2041 tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag
     2101 aatggcaaaa gcttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg
     2161 tcatcaaaat cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga
     2221 cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc
     2281 aggaacactg ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc
     2341 tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg
     2401 ataaaatgct tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc
     2461 tcatctgtaa catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca
     2521 tcgggcttcc catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc
     2581 catttatacc catataaatc agcatccatg ttggaattta atcgcggcct ggagcaagac
     2641 gtttcccgtt gaatatggct cataacaccc cttgtattac tgtttatgta agcagacagt
     2701 tttattgttc atgatgatat atttttatct tgtgcaatgt acatcagaga ttttgagaca
     2761 caaccaatta ttgaaggcct ccctaacggg gggccttttt ttgtttctgg tctcccgctt
     2821 aacgatcgtt ggctgtgttg acaattaatc atcggctcgt ataatgtgtg gaattgtgag
     2881 cgctcacaat tag
//
