LOCUS       sequence-408092-        2887 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  bm3R1 repressor plasmid for LPattB2 integration.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2887)
  AUTHORS   Triassi AJ, Fields BD, Monahan CE, Means JM, Park Y, Doosthosseini 
            H, Padmakumar JP, Isabella VM, Voigt CA
  TITLE     Redesign of an Escherichia coli Nissle treatment for phenylketonuria
            using insulated genomic landing pads and genetic circuits to reduce 
            burden.
  JOURNAL   Cell Syst. 2023 Jun 21;14(6):512-524.e12. doi: 
            10.1016/j.cels.2023.05.004.
  PUBMED    37348465
REFERENCE   2  (bases 1 to 2887)
  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"; doi: 
            "10.1016/j.cels.2023.05"; journalName: "Cell Syst"; date: 
            "2023-06-21- 21"; volume: "14"; issue: "6"; pages: "512-524.e12"
COMMENT     Sequence Label: pAJT251-bm3R1
FEATURES             Location/Qualifiers
     source          1..2887
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    785..804
                     /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."
     promoter        complement(join(810..816,817..832,833..838))
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    complement(928..975)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        1027..1118
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1119..1934
                     /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             1119..1934
                     /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     complement(1192..1211)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      1945..2039
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     oriT            2190..2298
                     /note="incP origin of transfer"
     rep_origin      2342..2730
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
ORIGIN
        1 gaatctcgcg tagacacata catgagcagc gaaaagaggt ggccaggggg atcacctggc
       61 agcatgctgc ctaataaaaa aggccggaaa atccggcctt ttgacgcttt agctctgacg
      121 gctcagtgct gcccacaggc tttcttcaac accggtcagc agttcatcgg tcaggctcag
      181 ataatcgttc tcgatcattt catacacttc cataaagctg ccaaacagaa ttgcaatcag
      241 tgcattttcc ggcagattac gaatcacacc ctgtttctga ccttcacgaa aaaaggtgca
      301 cacaaattca accagtttct gatatgccag acggctttct tcggtcagaa aggtgccctg
      361 gctatgggtt ttgataaaac ccagtgcacg cggatgattt ttggtaaagg tcaccatacc
      421 ttcaaaaatg tgatgaaagc catcacgata accatcacgt tcatttgcca gaccgctttc
      481 aatacactgc agaaattcat taacatgctg ctgaaacagt tcgttcacca ggctttcttt
      541 gtttttgaaa tagcgataaa tggtgcctgc accaactttt gcattttctg caatcatcgg
      601 catggtggtt gcatcaaaac cacgttctgc aaacagcagc aggcttgcgc taaaaattgc
      661 tttctgtttg gtcggggtgc tttccatcct cccggcctcg tttggttaaa caaaattatt
      721 tgtagaggac tgtttcggcc cttttgggcc atcgtcaggt cggatacaca tccggcgaca
      781 gtctaattgt gagcgctcac aattccacac attatacgag ccgatgatta attgtcaaca
      841 cagccaacga tcgttctccc agtacaatct gctctgatgc cctcgagtgc ggccgcacta
      901 gtgtcgacgg agctcgaatt cggatccgaa gttcctatac tttctagaga ataggaactt
      961 cggaatagga acttctattt ttataggtta atgtcatgat aataatggtt tcttagacca
     1021 tgcctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     1081 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagccatatt caacgggaaa
     1141 cgtcttgctc taggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat
     1201 gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgattgtat gggaagcccg
     1261 atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg
     1321 agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta
     1381 tccgtactcc tgatgatgca tggttactca ccactgcgat ccccgggaaa acagcattcc
     1441 aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc
     1501 tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgac cgcgtatttc
     1561 gtctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt gattttgatg
     1621 acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataaa cttttgccat
     1681 tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg
     1741 aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg
     1801 atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt
     1861 ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg
     1921 atgagttttt ctaaagctcg cttggactcc tgttgataga tccagtaatg acctcagaac
     1981 tccatctgga tttgttcaga acgctcggtt gccgccgggc gttttttatt ggtgagaatc
     2041 caagcttaag gtatactttc cgctgcataa ccctgcttcg gggtcattat agcgattttt
     2101 tcggtatatc catccttttt cgcacgatat acaggatttt gccaaagggt tcgtgtagac
     2161 tttccttggt gtatccaacg gcgtcagccg ggcaggatag gtgaagtagg cccacccgcg
     2221 agcgggtgtt ccttcttcac tgtcccttat tcgcacctgg cggtgctcaa cgggaatcct
     2281 gctctgcgag gctggccgat aagctacaag aagatccggc cacgatgcgt ccggcgtaga
     2341 ggatctgaag atcagcagtt caacctgttg atagtacgta ctaagctctc atgtttcacg
     2401 tactaagctc tcatgtttaa cgtactaagc tctcatgttt aacgaactaa accctcatgg
     2461 ctaacgtact aagctctcat ggctaacgta ctaagctctc atgtttcacg tactaagctc
     2521 tcatgtttga acaataaaat taatataaat cagcaactta aatagcctct aaggttttaa
     2581 gttttataag aaaaaaaaga atatataagg cttttaaagc ttttaaggtt taacggttgt
     2641 ggacaacaag ccagggatgt aacgcactga gaagccctta gagcctctca aagcaatttt
     2701 gagtgacaca ggaacactta acggctgaca tgggaattag ccatgggccc gtgcgaatca
     2761 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     2821 cctgattctg tggataaccg tattaccgcc tagccaagag cagtgccttg cagaagttct
     2881 cgggcga
//
