LOCUS       sequence-492178-        3440 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GoldenBraid2.0 Flp recombinase excisable antibiotic resistance 
            cassette in the grammar of an assembled transcription unit. May be 
            omega-assembled with another transcription unit..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3440)
  AUTHORS   Cooke MB, Welch KT, Deus Ramirez L, Wen AX, Marciano DC,, Herman C
  TITLE     GoldenBraid2.0 E. coli: A comprehensive and characterized toolkit 
            for enterics.
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3440)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pBSKa2_FRTspcFRT
FEATURES             Location/Qualifiers
     source          1..3440
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    44..77
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     promoter        158..249
                     /gene="bla"
                     /label=AmpR promoter
     CDS             250..1041
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     protein_bind    1054..1101
                     /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)."
     rep_origin      complement(1240..1828)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1320..1339)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        1943..2045
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             2046..2861
                     /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             2046..2861
                     /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(2119..2138)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2975..3430
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3062..3081)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3272..3293
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 cgtctcagtc aggagattcc ggggatccgt cgacctgcag ttcgaagttc ctattctcta
       61 gaaagtatag gaacttcaga gcgcttttga agctcacgtt caaatatgta tccgctcatg
      121 agacaataac cctggtagcg accgagtgag ctagctattt gtttattttt ctaaatacat
      181 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
      241 aggaagagta tgagggaagc ggtgatcgcc gaagtatcga ctcaactatc agaggtagtt
      301 ggcgtcatcg agcgccatct cgaaccgacg ttgctggccg tacatttgta cggctccgca
      361 gtggatggcg gcctgaagcc acacagtgat attgatttgc tggttacggt gaccgtaagg
      421 cttgatgaaa caacgcggcg agctttgatc aacgaccttt tggaaacttc ggcttcccct
      481 ggagagagcg agattctccg cgctgtagaa gtcaccattg ttgtgcacga cgacatcatt
      541 ccgtggcgtt atccagctaa gcgcgaactg caatttggag aatggcagcg caatgacatt
      601 cttgcaggta tcttcgagcc agccacgatc gacattgatc tggctatctt gctgacaaaa
      661 gcaagagaac atagcgttgc cttggtaggt ccagcggcgg aggaactctt tgatccggtt
      721 cctgaacagg atctatttga ggcgctaaat gaaaccttaa cgctatggaa ctcgccgccc
      781 gactgggctg gcgatgagcg aaatgtagtg cttacgttgt cccgcatttg gtacagcgca
      841 gtaaccggca aaatcgcgcc gaaggatgtc gctgccgact gggcaatgga gcgcctgccg
      901 gcccagtatc agcccgtcat acttgaagct agacaggctt atcttggaca agaagaagat
      961 cgcttggcct cgcgcgcaga tcagttggaa gaatttgtcc actacgtgaa aggcgagatc
     1021 accaaggtag tcggcaaata ataaggggat cttgaagttc ctattccgaa gttcctattc
     1081 tctagaaagt ataggaactt cgaagcagct ccagcctaca cgcttgagac gaagcttgac
     1141 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     1201 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     1261 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     1321 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     1381 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     1441 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     1501 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     1561 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     1621 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     1681 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     1741 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     1801 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     1861 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     1921 cttcacctag atccttttgt gatgatcggc acgtaagagg ttccaacttt caccataatg
     1981 aaataagatc actaccgggc gtattttttg agttatcgag attttcagga gctaaggaag
     2041 ctaaaatgag ccatattcaa cgggaaacgt cttgctcaag gccgcgatta aattccaaca
     2101 tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga
     2161 caatctaccg attgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag
     2221 gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg acggaattta
     2281 tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg ttactcacca
     2341 ctgcgatccc agggaaaaca gcattccagg tattagaaga atatcctgat tcaggtgaaa
     2401 atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct gtttgtaatt
     2461 gtccttttaa cagcgatcgc gtatttcgac tcgctcaggc gcaatcacga atgaataacg
     2521 gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct
     2581 ggaaagaaat gcataaactt ttgccattct caccggattc agtagtcact catggtgatt
     2641 tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt gatgttggac
     2701 gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc ctcggtgagt
     2761 tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga
     2821 ataaattgca gtttcatttg atgctcgatg agtttttctg atttttttta aggcagttat
     2881 tggtgccctt aaacgcctgg ttgctacgcc tgaataagtg ataataagcg gatgaatggc
     2941 agaaattcga tgataagctg tcaaacatga tgttacgcgc cctgtagcgg cgcattaagc
     3001 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     3061 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     3121 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     3181 aaacttgatt agggtgatgg ttcacgtagt gggccagcgc cctgatagac ggtttttcgc
     3241 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     3301 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     3361 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     3421 cttacaattt taacaagctt
//
