LOCUS       sequence-488058-        3772 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 3772)
  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  JOURNAL   Unpublished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 3772)
  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            "Unpublished"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: pBSKa1_FRTtetFRT
FEATURES             Location/Qualifiers
     source          1..3772
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    40..73
                     /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        join(104..109,110..126,127..132)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     CDS             180..1370
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     complement(532..551)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     protein_bind    1383..1430
                     /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(1573..2161)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1653..1672)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2276..2378
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             2379..3194
                     /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             2379..3194
                     /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(2452..2471)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3308..3763
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3395..3414)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3605..3626
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 cgtctcagga gattccgggg atccgtcgac ctgcagttcg aagttcctat tctctagaaa
       61 gtataggaac ttcagagcgc ttttgaagct cacgttctca tgtttgacag cttatcatcg
      121 ataagcttta atgcggtagt ttatcacagt taaattgcta acgcagtcag gcaccgtgta
      181 tgaaatctaa caatgcgctc atcgtcatcc tcggcaccgt caccctggat gctgtaggca
      241 taggcttggt tatgccggta ctgccgggcc tcttgcggga tatcgtccat tccgacagca
      301 tcgccagtca ctatggcgtg ctgctagcgc tatatgcgtt gatgcaattt ctatgcgcac
      361 ccgttctcgg agcactgtcc gaccgctttg gccgccgccc agtcctgctc gcttcgctac
      421 ttggagccac tatcgactac gcgatcatgg cgaccacacc cgtcctgtgg atcctctacg
      481 ccggacgcat cgtggccggc atcaccggcg ccacaggtgc ggttgctggc gcctatatcg
      541 ccgacatcac cgatggggaa gatcgggctc gccacttcgg gctcatgagc gcttgtttcg
      601 gcgtgggtat ggtggcaggc cccgtggccg ggggactgtt gggcgccatc tccttgcatg
      661 caccattcct tgcggcggcg gtgctcaacg gcctcaacct actactgggc tgcttcctaa
      721 tgcaggagtc gcataaggga gagcgtcgac cgatgccctt gagagccttc aacccagtca
      781 gctccttccg gtgggcgcgg ggcatgacta tcgtcgccgc acttatgact gtcttcttta
      841 tcatgcaact cgtaggacag gtgccggcag cgctctgggt cattttcggc gaggaccgct
      901 ttcgctggag cgcgacgatg atcggcctgt cgcttgcggt attcggaatc ttgcacgccc
      961 tcgctcaagc cttcgtcact ggtcccgcca ccaaacgttt cggcgagaag caggccatta
     1021 tcgccggcat ggcggccgac gcgctgggct acgtcttgct ggcgttcgcg acgcgaggct
     1081 ggatggcctt ccccattatg attcttctcg cttccggcgg catcgggatg cccgcgttgc
     1141 aggccatgct gtccaggcag gtagatgacg accatcaggg acagcttcaa ggatcgctcg
     1201 cggctcttac cagcctaact tcgatcattg gaccgctgat cgtcacggcg atttatgccg
     1261 cctcggcgag cacatggaac gggttggcat ggattgtagg cgccgcccta taccttgtct
     1321 gcctccccgc gttgcgtcgc ggtgcatgga gccgggccac ctcgacctga taaggggatc
     1381 ttgaagttcc tattccgaag ttcctattct ctagaaagta taggaacttc gaagcagctc
     1441 cagcctacac gctgtcatga gacgaattct gacacggtta tccacagaat caggggataa
     1501 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     1561 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     1621 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     1681 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     1741 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     1801 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     1861 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     1921 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     1981 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     2041 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     2101 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     2161 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     2221 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt tgtgatgatc
     2281 ggcacgtaag aggttccaac tttcaccata atgaaataag atcactaccg ggcgtatttt
     2341 ttgagttatc gagattttca ggagctaagg aagctaaaat gagccatatt caacgggaaa
     2401 cgtcttgctc aaggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat
     2461 gggctcgcga taatgtcggg caatcaggtg cgacaatcta ccgattgtat gggaagcccg
     2521 atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg
     2581 agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta
     2641 tccgtactcc tgatgatgca tggttactca ccactgcgat cccagggaaa acagcattcc
     2701 aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc
     2761 tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat cgcgtatttc
     2821 gactcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt gattttgatg
     2881 acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataaa cttttgccat
     2941 tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg
     3001 aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg
     3061 atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt
     3121 ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg
     3181 atgagttttt ctgatttttt ttaaggcagt tattggtgcc cttaaacgcc tggttgctac
     3241 gcctgaataa gtgataataa gcggatgaat ggcagaaatt cgatgataag ctgtcaaaca
     3301 tgatgttacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     3361 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     3421 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     3481 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     3541 agtgggccag cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     3601 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     3661 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     3721 aaatttaacg cgaattttaa caaaatatta acgcttacaa ttttaacgaa tt
//
