LOCUS       sequence-208064-        8272 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BTK assembled plasmid- broad-host-range GFP expression from PA1 
            promoter (spectinomycin resistance).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8272)
  AUTHORS   Leonard SP, Perutka J, Powell JE, Geng P, Richhart DD, Byrom M, Kar 
            S, Davies BW, Ellington AD, Moran NA, Barrick JE
  TITLE     Genetic Engineering of Bee Gut Microbiome Bacteria with a Toolkit 
            for Modular Assembly of Broad-Host-Range Plasmids.
  JOURNAL   ACS Synth Biol. 2018 May 18;7(5):1279-1290. doi: 
            10.1021/acssynbio.7b00399. Epub 2018 Apr 13.
  PUBMED    29608282
REFERENCE   2  (bases 1 to 8272)
  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: 
            "10.1021/acssynbio.7b00399"; journalName: "ACS Synth Biol"; date: 
            "2018-05-18- 18"; volume: "7"; issue: "5"; pages: "1279-1290"
COMMENT     Sequence Label: pBTK520
FEATURES             Location/Qualifiers
     source          1..8272
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(161..1012)
                     /codon_start=1
                     /gene="repC"
                     /product="replication protein C of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepC
                     /translation="MVKPKNKHSLSHVRHDPAHCLAPGLFRALKRGERKRSKLDVTYDY
                     GDGKRIEFSGPEPLGADDLRILQGLVAMAGPNGLVLGPEPKTEGGRQLRLFLEPKWEAV
                     TAECHVVKGSYRALAKEIGAEVDSGGALKHIQDCIERLWKVSIIAQNGRKRQGFRLLSE
                     YASDEADGRLYVALNPLIAQAVMGGGQHVRISMDEVRALDSETARLLHQRLCGWIDPGK
                     TGKASIDTLCGYVWPSEASGSTMRKRRKRVREALPELVALGWTVTEFAAGKYDITRPKA
                     AG"
     CDS             complement(999..1838)
                     /codon_start=1
                     /gene="repA"
                     /product="replication protein A of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepA
                     /translation="MATHKPINILEAFAAAPPPLDYVLPNMVAGTVGALVSPGGAGKSM
                     LALQLAAQIAGGPDLLEVGELPTGPVIYLPAEDPPTAIHHRLHALGAHLSAEERQAVAD
                     GLLIQPLIGSLPNIMAPEWFDGLKRAAEGRRLMVLDTLRRFHIEEENASGPMAQVIGRM
                     EAIAADTGCSIVFLHHASKGAAMMGAGDQQQASRGSSVLVDNIRWQSYLSSMTSAEAEE
                     WGVDDDQRRFFVRFGVSKANYGAPFADRWFRRHDGGVLKPAVLERQRKSKGVPRGEA"
     CDS             complement(2349..3320)
                     /codon_start=1
                     /gene="repB"
                     /product="replication protein B of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepB
                     /translation="MKNDRTLQAIGRQLKAMGCERFDIGVRDAPTGQMMNREWSAAEVL
                     QNTPWLKRMNAQGNDVYIRPAEQERHGLVLVDDLSEFDLDDMKAEGREPALVVETSPKN
                     YQAWVKVADAAGGELRGQIARTLASEYDADPASADSRHYGRLAGFTNRKDKHTTRAGYQ
                     PWVLLRESKGKTATAGPALVQQAGQQIEQAQRQQEKARRLASLELPERQLSRHRRTALD
                     EYRSEMAGLVKRFGHDLSKCDFIAAQKLASRGRSAEEIGKAMAEASPALAERKPGHEAD
                     YIERTVSKVMGLPSVQLARAELARAPAPRQRGMDRGGPDFSM"
     oriT            4559..4646
                     /label=RSF1010 oriT
                     /note="origin of transfer of the broad-host-range plasmid 
                     RSF1010 (Scholz et al., 1989)"
     rep_origin      4988..5382
                     /label=RSF1010 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     RSF1010; requires the RSF1010 RepA/B/C proteins for 
                     replication (Scholz et al., 1989)"
     protein_bind    6089..6113
                     /label=lac operator
                     /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)."
     terminator      7129..7200
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      7216..7243
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(7262..8053)
                     /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"
ORIGIN
        1 acgctacttg cattacagct tacgaaccga acaggcttat gtccactggg ttcgtgcctt
       61 catccgtttc cacggtgtgc gtcacccggc aaccttgggt agcagcgaag tcgaggcatt
      121 tctgtcctgg ctggtcatga ccaaaatccc ttaacgtgag tcagcctgcc gccttgggcc
      181 gggtgatgtc gtacttgccc gccgcgaact cggttaccgt ccagcccagc gcgaccagct
      241 ccggcaacgc ctcgcgcacc cgcttgcggc gcttgcgcat ggtcgaacca ctggcctctg
      301 acggccagac atagccgcac aaggtatcta tggaagcctt gccggttttg ccggggtcga
      361 tccagccaca cagccgctgg tgcagcaggc gggcggtttc gctgtccagc gcccgcacct
      421 cgtccatgct gatgcgcaca tgctggccgc cacccatgac ggcctgcgcg atcaaggggt
      481 tcagggccac gtacaggcgc ccgtccgcct cgtcgctggc gtactccgac agcagccgaa
      541 acccctgccg cttgcggcca ttctgggcga tgatggatac cttccaaagg cgctcgatgc
      601 agtcctgtat gtgcttgagc gccccaccac tatcgacctc tgccccgatt tcctttgcca
      661 gcgcccgata gctacctttg accacatggc attcagcggt gacggcctcc cacttgggtt
      721 ccaggaacag ccggagctgc cgtccgcctt cggtcttggg ttccgggcca agcactaggc
      781 cattaggccc agccatggcc accagccctt gcaggatgcg cagatcatca gcgcccagcg
      841 gctccgggcc gctgaactcg atccgcttgc cgtcgccgta gtcatacgtc acgtccagct
      901 tgctgcgctt gcgctcgccc cgcttgaggg cacggaacag gccgggggcc agacagtgcg
      961 ccgggtcgtg ccggacgtgg ctgaggctgt gcttgttctt aggcttcacc acggggcacc
     1021 cccttgctct tgcgctgcct ctccagcacg gcgggcttga gcaccccgcc gtcatgccgc
     1081 ctgaaccacc gatcagcgaa cggtgcgcca tagttggcct tgctcacacc gaagcggacg
     1141 aagaaccggc gctggtcgtc gtccacaccc cattcctcgg cctcggcgct ggtcatgctc
     1201 gacaggtagg actgccagcg gatgttatcg accagtaccg agctgccccg gctggcctgc
     1261 tgctggtcgc ctgcgcccat catggccgcg cccttgctgg catggtgcag gaacacgata
     1321 gagcacccgg tatcggcggc gatggcctcc atgcgaccga tgacctgggc catggggccg
     1381 ctggcgtttt cttcctcgat gtggaaccgg cgcagcgtgt ccagcaccat caggcggcgg
     1441 ccctcggcgg cgcgcttgag gccgtcgaac cactccgggg ccatgatgtt gggcaggctg
     1501 ccgatcagcg gctggatcag caggccgtca gccacggctt gccgttcctc ggcgctgagg
     1561 tgcgccccaa gggcgtgcag gcggtgatga atggcggtgg gcgggtcttc ggcgggcagg
     1621 tagatcaccg ggccggtggg cagttcgccc acctccagca gatccggccc gcctgcaatc
     1681 tgtgcggcca gttgcagggc cagcatggat ttaccggcac caccgggcga caccagcgcc
     1741 ccgaccgtac cggccaccat gttgggcaaa acgtagtcca gcggtggcgg cgctgctgcg
     1801 aacgcctcca gaatattgat aggcttatgg gtagccattg attgcctcct ttgcaggcag
     1861 ttggtggtta ggcgctggcg gggtcactac ccccgccctg cgccgctctg agttcttcca
     1921 ggcactcgcg cagcgcctcg tattcgtcgt cggtcagcca gaacttgcgc tgacgcatcc
     1981 ctttggcctt catgcgctcg gcatatcgcg cttggcgtac agcgtcaggg ctggccagca
     2041 ggtcgccggt ctgcttgtcc ttttggtctt tcatatcagt caccgagaaa cttgccgggg
     2101 ccgaaaggct tgtcttcgcg gaacaaggac aaggtgcagc cgtcaaggtt aaggctggcc
     2161 atatcagcga ctgaaaagcg gccagcctcg gccttgtttg acgtataacc aaagccaccg
     2221 ggcaaccaat agcccttgtc acttttgatc aggtagaccg accctgaagc gcttttttcg
     2281 tattccataa aacccccttc tgtgcgtgag tactcatagt ataacaggcg tgagtaccaa
     2341 cgcaagcact acatgctgaa atctggcccg cccctgtcca tgcctcgctg gcggggtgcc
     2401 ggtgcccgtg ccagctcggc ccgcgcaagc tggacgctgg gcagacccat gaccttgctg
     2461 acggtgcgct cgatgtaatc cgcttcgtgg ccgggcttgc gctctgccag cgctgggctg
     2521 gcctcggcca tggccttgcc gatttcctcg gcactgcggc cccggctggc cagcttctgc
     2581 gcggcgataa agtcgcactt gctgaggtca tgaccgaagc gcttgaccag cccggccatc
     2641 tcgctgcggt actcgtccag cgccgtgcgc cggtggcggc taagctgccg ctcgggcagt
     2701 tcgaggctgg ccagcctgcg ggccttctcc tgctgccgct gggcctgctc gatctgctgg
     2761 ccagcctgct gcaccagcgc cgggccagcg gtggcggtct tgcccttgga ttcacgcagc
     2821 agcacccacg gctgataacc ggcgcgggtg gtgtgcttgt ccttgcggtt ggtgaagccc
     2881 gccaagcggc catagtggcg gctgtcggcg ctggccgggt cggcgtcgta ctcgctggcc
     2941 agcgtccggg caatctgccc ccgaagttca ccgcctgcgg cgtcggccac cttgacccat
     3001 gcctgatagt tcttcgggct ggtttccact accagggcag gctcccggcc ctcggctttc
     3061 atgtcatcca ggtcaaactc gctgaggtcg tccaccagca ccagaccatg ccgctcctgc
     3121 tcggcgggcc tgatatacac gtcattgccc tgggcattca tccgcttgag ccatggcgtg
     3181 ttctggagca cttcggcggc tgaccattcc cggttcatca tctggccggt gggtgcgtcc
     3241 ctgacgccga tatcgaagcg ctcacagccc atggccttga gctgtcggcc tatggcctgc
     3301 aaagtcctgt cgttcttcat cgggccacca agcgcagcca gatcgagccg tcctcggttg
     3361 tcagtggcgt caggtcgagc aagagcaacg atgcgatcag cagcaccacc gtaggcatca
     3421 tggaagccag catcacggtt agccatagct tccagtgcca cccccgcgac gcgctccggg
     3481 cgctctgcgc ggcgctgctc acctcggcgg ctacctcccg caactctttg gccagctcca
     3541 cccatgccgc ccctgtctgg cgctgggctt tcagccactc cgccgcctgc gcctcgctgg
     3601 cctgcttggt ctggctcatg acctgccggg cttcgtcggc cagtgtcgcc atgctctggg
     3661 ccagcggttc gatctgctcc gctaactcgt tgatgcctct ggatttcttc actctgtcga
     3721 ttgcgttcat ggtctattgc ctcccggtat tcctgtaagt cgatgatctg ggcgttggcg
     3781 gtgtcgatgt tcagggccac gtctgcccgg tcggtgcgga tgccccggcc ttccatctcc
     3841 accacgttcg gccccaggtg aacaccgggc aggcgctcga tgccctgcgc ctcaagtgtt
     3901 ctgtggtcaa tgcgggcgtc gtggccagcc cgctctaatg cccggttggc atggtcggcc
     3961 catgcctcgc gggtctgctc aagccatgcc ttgggcttga gcgcttcggt cttctgtgcc
     4021 ccgcccttct ccggggtctt gccgttgtac cgcttgaacc actgagcggc gggccgctcg
     4081 atgccgtcat tgatccgctc ggagatcatc aggtggcagt gcgggttctc gccgccaccg
     4141 gcatggatgg ccagcgtata cggcaggcgc tcggcaccgg tcaggtgctg ggcgaactcg
     4201 gacgccagcg ccttctgctg gtcgagggtc agctcgaccg gcagggcaaa ttcgacctcc
     4261 ttgaacagcc gcccattggc gcgttcatac aggtcggcag catcccagta gtcggcgggc
     4321 cgctcgacga actccggcat gtgcccggat tcggcgtgca agacttcatc catgtcgcgg
     4381 gcatacttgc cttcgcgctg gatgtagtcg gccttggccc tggccgattg gccgcccgac
     4441 ctgctgccgg ttttcgccgt aaggtgataa atcgccatgc tgcctcgctg ttgcttttgc
     4501 ttttcggctc catgcaatgg ccctcggaga gcgcaccgcc cgaagggtgg ccgttaggcc
     4561 agtttctcga agagaaaccg gtaagtgcgc cctcccctac aaagtagggt cgggattgcc
     4621 gccgctgtgc ctccatgata gcctacgaga cagcacatta acaatggggt gtcaagatgg
     4681 ttaaggggag caacaaggcg gcggatcggc tggccaagct cgaagaacaa cgagcgcgaa
     4741 tcaatgccga aattcagcgg gagcgggcaa gggaacagca gcaagagcgc aagaacgaaa
     4801 caaggcgcaa ggtgctggtg ggggccatga ttttggccaa ggtgaacagc agcgagtggc
     4861 cggaggatcg gctcatggcg gcaatggatg cgtaccttga acgcgaccac gaccgcgcct
     4921 tgttcggtct gccgccacgc cagaaggatg agccgggctg aatgatcgac cgagacaggc
     4981 cctgcggggc tgcacacgcg cccccaccct tcgggtaggg ggaaaggccg ctaaagcggc
     5041 taaaagcgct ccagcgtatt tctgcggggt ttggtgtggg gtttagcggg ctttgcccgc
     5101 ctttccccct gccgcgcagc ggtggggcgg tgtgtagcct agcgcagcga atagaccagc
     5161 tatccggcct ctggccgggc atattgggca agggcagcag cgccccacaa gggcgctgat
     5221 aaccgcgcct agtggattat tcttagataa tcatggatgg atttttccaa caccccgcca
     5281 gcccccgccc ctgctgggtt tgcaggtttg ggggcgtgac agttattgca ggggttcgtg
     5341 acagttattg caggggggcg tgacagttat tgcaggggtt cgtgacagtt agtacgggag
     5401 tgacgggcac tggctggcaa tgtctagcaa cggcaggcat ttcggctgag ggtaaaagaa
     5461 ctttccgcta agcgatagac tgtatgtaaa cacagtattg caaggacgcg gaacatgcct
     5521 catgtggcgg ccaggacggc cagccgggat cgggatactg gtcgttacca gagccaccga
     5581 cccgagcaaa cccttctcta tcagatcgtt gacgagtatt acccggcatt cgctgcgctt
     5641 atggcagagc agggaaagga attgccgggc tatgtgcaac gggaatttga agaatttctc
     5701 caatgcgggc ggctggagca tggctttcta cgggttcgct gcgagtcttg ccacgccgag
     5761 cacctggtcg ctttcagaaa tcaatctaaa gtatatatga gtaaacttgg tctgacaggc
     5821 ccctgaattc gcatctagat ggtagagcca caaacagccg gtacaagcaa cgatctccag
     5881 gaccatctga atcatgcgcg gatgacacga actcacgacg gcgatcacag acattaaccc
     5941 acagtacaga cactgcgaca acgtggcaat tcgtcgcaat accgtctcac tgaactggcc
     6001 gataattgca gacgaacgtt atcaaaaaga gtattgactt aaagtctaac ctataggata
     6061 cttacagcca tcgagaggga cacggcgagg aattgtgagc ggataacaat tccatacaga
     6121 aacagaggag atattacata tgagtaaagg agaagagctt ttcacaggag ttgtcccaat
     6181 cctcgtggaa ttagacggtg atgttaatgg gcacaagttc tctgtcagtg gagagggtga
     6241 aggcgacgca acatatggca agctgaccct taaatttatt tgcaccacgg gtaaactacc
     6301 tgttccatgg ccaacactgg tcactacgtt cgggtatggg gttcagtgct ttgcgcgcta
     6361 cccagatcac atgaaacagc acgacttttt caagagtgca atgcccgaag gctatgtaca
     6421 ggagagaacc atctttttta aggatgacgg caactataag acacgcgccg aagtgaagtt
     6481 cgagggtgat acccttgtta atagaatcga gttaaagggt attgacttta aggaagatgg
     6541 aaatatttta ggccacaaac tggaatataa ctataactcc cataatgtgt acattatggc
     6601 cgacaagcaa aagaacggta tcaaggttaa cttcaagatc agacacaaca ttgaggatgg
     6661 aagcgttcaa ctagccgacc attaccaaca aaacacccca attggcgatg ggcctgtgct
     6721 gttaccagac aaccattacc tgtccactca atctgccctt tcgaaagatc ccaacgaaaa
     6781 gcgcgaccac atggtccttc ttgagtttgt cacggctgct gggattacac acggcatgga
     6841 tgaactatac aaataaatcc gtaatcgtta atccgcaaat aacgtaaaaa cccgcttcgg
     6901 cgggtttttt tatgggggga gtttagggaa agagcatttg tcagctgcca atgagacgac
     6961 ggggtcatca cggctcatca tgcgccaaac aaatgtgtgc aatacacgct cggatgactg
     7021 catgatgacc gcactgactg gggacagcag atccacctaa gcctgtgaga gaagcagaca
     7081 cccgacagat caaggcagtt aactagtgca ctgcagtaca ccaggcatca aataaaacga
     7141 aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc
     7201 tactagagtc acactggctc accttcgggt gggcctttct gcgtttatac cgagcggccg
     7261 cttatttgcc gactaccttg gtgatctcgc ctttcacgta gtggacaaat tcttccaact
     7321 gatctgcgcg cgaggccaag cgatcttctt cttgtccaag ataagcctgt ctagcttcaa
     7381 gtatgacggg ctgatactgg gccggcaggc gctccattgc ccagtcggca gcgacatcct
     7441 tcggcgcgat tttgccggtt actgcgctgt accaaatgcg ggacaacgta agcactacat
     7501 ttcgctcatc gccagcccag tcgggcggcg agttccatag cgttaaggtt tcatttagcg
     7561 cctcaaatag atcctgttca ggaaccggat caaagagttc ctccgccgct ggacctacca
     7621 aggcaacgct atgttctctt gcttttgtca gcaagatagc cagatcaatg tcgatcgtgg
     7681 ctggctcgaa gatacccgca agaatgtcat tgcgctgcca ttctccaaat tgcagttcgc
     7741 gcttagctgg ataacgccac ggaatgatgt cgtcgtgcac aacaatggtg acttctacag
     7801 cgcggagaat ctcgctctct ccaggggaag ccgaagtttc caaaaggtcg ttgatcaaag
     7861 ctcgccgcgt tgtttcatca agccttacgg tcaccgtaac cagcaaatca atatcactgt
     7921 gtggcttcag gccgccatcc actgcggagc cgtacaaatg tacggccagc aacgtcggtt
     7981 cgagatggcg ctcgatgacg ccaactacct ctgatagttg agtcgatact tcggcgatca
     8041 ccgcttccct cataatgttt aactttgttt tagggcgact gccctgctgc gtaacatcgt
     8101 tgctgctcca taacatcaaa catcgaccca cggcgtaacg cgcttgctgc ttggatgccc
     8161 gaggcataga ctgtacccca aaaaaacagt cataacaagc catgaaaacc gccactgcgc
     8221 cgttaccacc gctgcgttcg gtcaaggttc tggaccagtt gcgtgagcgc at
//
