LOCUS       sequence-503365-        5373 bp    DNA     circular SYN 18-MAR-2026
DEFINITION  Cumate-inducible Cre recombinase; cumate activates P_cymRC to 
            express cre[LVA], which excises loxP-flanked transcriptional 
            terminators and activates eyfp. Use with Marionette Sensor 
            Collection strains.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5373)
  AUTHORS   Garabello E, Yoon H, Reid MC, Giometto A
  TITLE     Tunable Low-Rate Genomic Recombination with Cre-lox in Escherichia 
            coli : A Versatile Tool for Environmental Biosensing and Synthetic 
            Biology.
  JOURNAL   bioRxiv [Preprint]. 2025 Sep 2:2024.10.02.616356. doi: 
            10.1101/2024.10.02.616356.
  PUBMED    40949998
REFERENCE   2  (bases 1 to 5373)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Mar 18, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2025 Sep 2:2024.10.02.616356. doi: 
            10.1101/2024.10.02.616356."
COMMENT     Sequence Label: p-cymR-cre[LVA]-loxPP-eyfp
FEATURES             Location/Qualifiers
     source          1..5373
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(399..1214)
                     /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(399..1214)
                     /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     1122..1141
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1206..1225
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     protein_bind    1387..1414
                     /label=CuO
                     /bound_moiety="CymR"
                     /note="CymR-binding P2 operator sequence from the p-cmt 
                     operon of Pseudomonas putida (Mullick et al., 2006)"
     protein_bind    1445..1472
                     /label=CuO
                     /bound_moiety="CymR"
                     /note="CymR-binding P2 operator sequence from the p-cmt 
                     operon of Pseudomonas putida (Mullick et al., 2006)"
     CDS             2712..2723
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     protein_bind    complement(2828..2861)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(2978..3011)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             join(3084..3086,3087..3089,3090..3803)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3129..3150)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3390..3409)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3737..3758
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4754..4773)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     rep_origin      complement(join(5138..5373,1..310))
                     /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(5218..5237)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgcacgaac cccccgttca gtccgaccgc tgcgccttat ccggtaacta tcgtcttgag
       61 tccaacccgg aaagacatgc aaaagcacca ctggcagcag ccactggtaa ttgatttaga
      121 ggagttagtc ttgaagtcat gcgccggtta aggctaaact gaaaggacaa gttttggtga
      181 ctgcgctcct ccaagccagt tacctcggtt caaagagttg gtagctcaga gaaccttcga
      241 aaaaccgccc tgcaaggcgg ttttttcgtt ttcagagcaa gagattacgc gcagaccaaa
      301 acgatctcaa gaagatcatc ttattaaggg gtctgacgct cagtggaacg aaaaatcaat
      361 ctaaagtata tatgagtaaa cttggtctga cagttacctt agaaaaactc atcgagcatc
      421 aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt
      481 ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat
      541 cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa
      601 ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa
      661 agcttatgca tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa
      721 tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg
      781 cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact
      841 gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct
      901 gttttcccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc
      961 ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta
     1021 acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc
     1081 ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac
     1141 ccatataaat cagcatccat gttggaattt aatcgcggcc tggagcaaga cgtttcccgt
     1201 tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt
     1261 catgatgata tatttttatc ttgtgcaatg tacatcagag attttgagac acaaccaatt
     1321 attgaaggcc tccctaacgg ggggcctttt tttgtttctg gtctcccgct taacgatcgt
     1381 tggctgaaca aacagacaat ctggtctgtt tgtattatgg aaaatttttc tgtataatag
     1441 attcaacaaa cagacaatct ggtctgtttg tattatagct gtcaccggat gtgctttccg
     1501 gtctgatgaa agaggggaaa tactagatgt ccaacctgct gacggtgcac cagaatctgc
     1561 cggcactgcc ggtcgatgca accagtgacg aagtgcgcaa aaatctgatg gatatgtttc
     1621 gtgaccgcca agccttcagc gaacatacgt ggaaaatgct gctgtcggtt tgccgtagct
     1681 gggcggcctg gtgtaaactg aacaatcgca aatggtttcc ggcagaaccg gaagatgtgc
     1741 gtgactatct gctgtacctg caggcacgtg gtctggcagt taaaaccatc cagcaacatc
     1801 tgggccaact gaacatgctg caccgtcgct ctggtctgcc gcgtccgagt gattccaatg
     1861 ccgtcagtct ggtgatgcgt cgcattcgta aagaaaacgt ggatgcaggc gaacgcgcta
     1921 aacaggcact ggcttttgaa cgtaccgatt tcgaccaagt tcgctctctg atggaaaaca
     1981 gtgatcgttg ccaggacatc cgcaatctgg cattcctggg tattgcttat aacaccctgc
     2041 tgcgcattgc agaaatcgct cgtattcgcg tgaaagatat cagccgtacg gacggcggtc
     2101 gcatgctgat tcacatcggc cgtaccaaaa cgctggtttc caccgcgggc gtcgaaaaag
     2161 ccctgtcact gggtgtcacg aaactggtgg aacgctggat ttcagtttcg ggcgtcgcag
     2221 atgacccgaa caattacctg ttttgtcgtg tgcgcaaaaa tggtgttgca gctccgagcg
     2281 ctacctctca gctgagtacg cgtgcgctgg aaggcatctt cgaagccacc catcgcctga
     2341 tttatggcgc gaaagatgac agcggtcagc gttacctggc atggtccggt cactcagctc
     2401 gtgttggtgc agcacgtgat atggcacgtg caggtgtctc tatcccggaa attatgcagg
     2461 ccggcggttg gacgaacgtg aatattgtta tgaactatat tcgtaacctg gactctgaaa
     2521 cgggtgcgat ggtgcgtctg ctggaagatg gcgacaggcc tgcagcaaac gacgaaaact
     2581 acgctttagt agcttaagct ttgatcaatg acgaacaata aggcctccct aacggggggc
     2641 cttttttatt gataacaaaa cacagataaa aaaaatcctt agctttcgct aaggatgatt
     2701 tctatccaat tattgaaggc cgctaacgcg gccttttttt gtttctggtc tcccgattat
     2761 caaaaagagt attgcattaa agtctaacct ataggaatct tacagccatc gagagggaca
     2821 cggcgaaata acttcgtata gcatacatta tacgaagtta tcaaacgcat gagaaagccc
     2881 ccggaagatc accttccggg ggctttttta ttgcgcacct cggtaccaaa ttccagaaaa
     2941 gagacgcttt cgagcgtctt ttttcgtttt ggtccacata acttcgtata gcatacatta
     3001 tacgaagtta ttttaagaag gaaggacgaa acagcctcta caaataattt tgtttaatac
     3061 tagagaaaga ggggaaatac tagatggtga gcaagggcga ggagctgttc accggggtgg
     3121 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     3181 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accacaggca
     3241 agctgcccgt gccctggccc accctcgtga ccaccttcgg ctacggcctg caatgcttcg
     3301 cccgctaccc cgaccacatg aagctgcacg acttcttcaa gtccgccatg cccgaaggct
     3361 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     3421 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     3481 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     3541 tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg
     3601 aggacggcag cgtgcagctc gccgaccact accagcagaa caccccaatc ggcgacggcc
     3661 ccgtgctgct gcccgacaac cactacctta gctaccagtc cgccctgagc aaagacccca
     3721 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     3781 gcatggacga gctgtacaag taactcggta ccaaattcca gaaaagaggc ctcccgaaag
     3841 gggggccttt tttcgttttg gtccaatggc ggcgcgccat cgaatggtgc aaaacctttc
     3901 gcggtatggc atgatagcgc ccggaagaga gtcaattcag ggtggtgaat atgagcccga
     3961 aacgtcgtac ccaggcagaa cgtgcaatgg aaacccaggg taaactgatt gcagcagcac
     4021 tgggtgttct gcgtgaaaaa ggttatgcag gttttcgtat tgcagatgtt ccgggtgcag
     4081 ccggtgttag ccgtggtgca cagagccatc attttccgac caaactggaa ctgctgctgg
     4141 caacctttga atggctgtat gagcagatta ccgaacgtag ccgtgcacgt ctggcaaaac
     4201 tgaaaccgga agatgatgtt attcagcaga tgctggatga tgcagcagaa ttttttctgg
     4261 atgatgattt tagcatcggc ctggatctga ttgttgcagc agatcgtgat ccggcactgc
     4321 gtgaaggtat tcagcgtacc gttgaacgta atcgttttgt tgttgaagat atgtggctgg
     4381 gtgtgctggt gagccgtggt ctgagccgtg atgatgccga agatattctg tggctgattt
     4441 ttaacagcgt tcgtggtctg gtagttcgta gcctgtggca gaaagataaa gaacgttttg
     4501 aacgtgtgcg taatagcacc ctggaaattg cacgtgaacg ttatgcaaaa ttcaaacgtt
     4561 gataaggatc ctaattggta acgaatcaga caattgacgg ctcgagggag tagcataggg
     4621 tttgcagaat ccctgcttcg tccatttgac aggcacatta tgcatcgatg ataagctgtc
     4681 aaacatgagc agatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt
     4741 gcggttgctg gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc
     4801 gggctcatga gcaaatattt tatctgaggt gcttcctcgc tcactgactc gctgcacgag
     4861 gcagacctca gcgctagcgg agtgtatact ggcttactat gttggcactg atgagggtgt
     4921 cagtgaagtg cttcatgtgg caggagaaaa aaggctgcac cggtgcgtca gcagaatatg
     4981 tgatacagga tatattccgc ttcctcgctc actgactcgc tacgctcggt cgttcgactg
     5041 cggcgagcgg aaatggctta cgaacggggc ggagatttcc tggaagatgc caggaagata
     5101 cttaacaggg aagtgagagg gccgcggcaa agccgttttt ccataggctc cgcccccctg
     5161 acaagcatca cgaaatctga cgctcaaatc agtggtggcg aaacccgaca ggactataaa
     5221 gataccaggc gtttccccct ggcggctccc tcgtgcgctc tcctgttcct gcctttcggt
     5281 ttaccggtgt cattccgctg ttatggccgc gtttgtctca ttccacgcct gacactcagt
     5341 tccgggtagg cagttcgctc caagctggac tgt
//
