LOCUS       sequence-208066-        7439 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BTK Assembled plasmid- ready for Stage 2 assembly (ConLS' and ConRE'
            connectors with spectinomycin resistance).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7439)
  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 7439)
  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: pBTK527a
FEATURES             Location/Qualifiers
     source          1..7439
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(253..1224)
                     /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            2463..2550
                     /label=RSF1010 oriT
                     /note="origin of transfer of the broad-host-range plasmid 
                     RSF1010 (Scholz et al., 1989)"
     rep_origin      2892..3286
                     /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)"
     CDS             complement(4333..5124)
                     /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"
     CDS             complement(5504..6355)
                     /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(6342..7181)
                     /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"
ORIGIN
        1 ggggccgaaa ggcttgtctt cgcggaacaa ggacaaggtg cagccgtcaa ggttaaggct
       61 ggccatatca gcgactgaaa agcggccagc ctcggccttg tttgacgtat aaccaaagcc
      121 accgggcaac caatagccct tgtcactttt gatcaggtag accgaccctg aagcgctttt
      181 ttcgtattcc ataaaacccc cttctgtgcg tgagtactca tagtataaca ggcgtgagta
      241 ccaacgcaag cactacatgc tgaaatctgg cccgcccctg tccatgcctc gctggcgggg
      301 tgccggtgcc cgtgccagct cggcccgcgc aagctggacg ctgggcagac ccatgacctt
      361 gctgacggtg cgctcgatgt aatccgcttc gtggccgggc ttgcgctctg ccagcgctgg
      421 gctggcctcg gccatggcct tgccgatttc ctcggcactg cggccccggc tggccagctt
      481 ctgcgcggcg ataaagtcgc acttgctgag gtcatgaccg aagcgcttga ccagcccggc
      541 catctcgctg cggtactcgt ccagcgccgt gcgccggtgg cggctaagct gccgctcggg
      601 cagttcgagg ctggccagcc tgcgggcctt ctcctgctgc cgctgggcct gctcgatctg
      661 ctggccagcc tgctgcacca gcgccgggcc agcggtggcg gtcttgccct tggattcacg
      721 cagcagcacc cacggctgat aaccggcgcg ggtggtgtgc ttgtccttgc ggttggtgaa
      781 gcccgccaag cggccatagt ggcggctgtc ggcgctggcc gggtcggcgt cgtactcgct
      841 ggccagcgtc cgggcaatct gcccccgaag ttcaccgcct gcggcgtcgg ccaccttgac
      901 ccatgcctga tagttcttcg ggctggtttc cactaccagg gcaggctccc ggccctcggc
      961 tttcatgtca tccaggtcaa actcgctgag gtcgtccacc agcaccagac catgccgctc
     1021 ctgctcggcg ggcctgatat acacgtcatt gccctgggca ttcatccgct tgagccatgg
     1081 cgtgttctgg agcacttcgg cggctgacca ttcccggttc atcatctggc cggtgggtgc
     1141 gtccctgacg ccgatatcga agcgctcaca gcccatggcc ttgagctgtc ggcctatggc
     1201 ctgcaaagtc ctgtcgttct tcatcgggcc accaagcgca gccagatcga gccgtcctcg
     1261 gttgtcagtg gcgtcaggtc gagcaagagc aacgatgcga tcagcagcac caccgtaggc
     1321 atcatggaag ccagcatcac ggttagccat agcttccagt gccacccccg cgacgcgctc
     1381 cgggcgctct gcgcggcgct gctcacctcg gcggctacct cccgcaactc tttggccagc
     1441 tccacccatg ccgcccctgt ctggcgctgg gctttcagcc actccgccgc ctgcgcctcg
     1501 ctggcctgct tggtctggct catgacctgc cgggcttcgt cggccagtgt cgccatgctc
     1561 tgggccagcg gttcgatctg ctccgctaac tcgttgatgc ctctggattt cttcactctg
     1621 tcgattgcgt tcatggtcta ttgcctcccg gtattcctgt aagtcgatga tctgggcgtt
     1681 ggcggtgtcg atgttcaggg ccacgtctgc ccggtcggtg cggatgcccc ggccttccat
     1741 ctccaccacg ttcggcccca ggtgaacacc gggcaggcgc tcgatgccct gcgcctcaag
     1801 tgttctgtgg tcaatgcggg cgtcgtggcc agcccgctct aatgcccggt tggcatggtc
     1861 ggcccatgcc tcgcgggtct gctcaagcca tgccttgggc ttgagcgctt cggtcttctg
     1921 tgccccgccc ttctccgggg tcttgccgtt gtaccgcttg aaccactgag cggcgggccg
     1981 ctcgatgccg tcattgatcc gctcggagat catcaggtgg cagtgcgggt tctcgccgcc
     2041 accggcatgg atggccagcg tatacggcag gcgctcggca ccggtcaggt gctgggcgaa
     2101 ctcggacgcc agcgccttct gctggtcgag ggtcagctcg accggcaggg caaattcgac
     2161 ctccttgaac agccgcccat tggcgcgttc atacaggtcg gcagcatccc agtagtcggc
     2221 gggccgctcg acgaactccg gcatgtgccc ggattcggcg tgcaagactt catccatgtc
     2281 gcgggcatac ttgccttcgc gctggatgta gtcggccttg gccctggccg attggccgcc
     2341 cgacctgctg ccggttttcg ccgtaaggtg ataaatcgcc atgctgcctc gctgttgctt
     2401 ttgcttttcg gctccatgca atggccctcg gagagcgcac cgcccgaagg gtggccgtta
     2461 ggccagtttc tcgaagagaa accggtaagt gcgccctccc ctacaaagta gggtcgggat
     2521 tgccgccgct gtgcctccat gatagcctac gagacagcac attaacaatg gggtgtcaag
     2581 atggttaagg ggagcaacaa ggcggcggat cggctggcca agctcgaaga acaacgagcg
     2641 cgaatcaatg ccgaaattca gcgggagcgg gcaagggaac agcagcaaga gcgcaagaac
     2701 gaaacaaggc gcaaggtgct ggtgggggcc atgattttgg ccaaggtgaa cagcagcgag
     2761 tggccggagg atcggctcat ggcggcaatg gatgcgtacc ttgaacgcga ccacgaccgc
     2821 gccttgttcg gtctgccgcc acgccagaag gatgagccgg gctgaatgat cgaccgagac
     2881 aggccctgcg gggctgcaca cgcgccccca cccttcgggt agggggaaag gccgctaaag
     2941 cggctaaaag cgctccagcg tatttctgcg gggtttggtg tggggtttag cgggctttgc
     3001 ccgcctttcc ccctgccgcg cagcggtggg gcggtgtgta gcctagcgca gcgaatagac
     3061 cagctatccg gcctctggcc gggcatattg ggcaagggca gcagcgcccc acaagggcgc
     3121 tgataaccgc gcctagtgga ttattcttag ataatcatgg atggattttt ccaacacccc
     3181 gccagccccc gcccctgctg ggtttgcagg tttgggggcg tgacagttat tgcaggggtt
     3241 cgtgacagtt attgcagggg ggcgtgacag ttattgcagg ggttcgtgac agttagtacg
     3301 ggagtgacgg gcactggctg gcaatgtcta gcaacggcag gcatttcggc tgagggtaaa
     3361 agaactttcc gctaagcgat agactgtatg taaacacagt attgcaagga cgcggaacat
     3421 gcctcatgtg gcggccagga cggccagccg ggatcgggat actggtcgtt accagagcca
     3481 ccgacccgag caaacccttc tctatcagat cgttgacgag tattacccgg cattcgctgc
     3541 gcttatggca gagcagggaa aggaattgcc gggctatgtg caacgggaat ttgaagaatt
     3601 tctccaatgc gggcggctgg agcatggctt tctacgggtt cgctgcgagt cttgccacgc
     3661 cgagcacctg gtcgctttca gaaatcaatc taaagtatat atgagtaaac ttggtctgac
     3721 aggcccctga attcgcatct agactgatga gacgtggtag agccacaaac agccggtaca
     3781 agcaacgatc tccaggacca tctgaatcat gcgcggatga cacgaactca cgacggcgat
     3841 cacagacatt aacccacagt acagacactg cgacaacgtg gcaattcgtc gcaatacaac
     3901 gtaacgggct tcccatacaa tcgattagag attgtcatcg cacctgattg cccgacatta
     3961 tcgcgagtta taacccattt atacccatat aaatcagcat ccatgttgga atttaatcgc
     4021 gtgagcctat gtaacgggct tcccatacaa tcgattagag attgtcatcg cacctgattg
     4081 cccgacatta tcgcgagtta taacccattt atacccatat aaatcagcat ccatgttgga
     4141 atttaatcgc gtgagccatc cgtaatcgtt aatccgcaaa taacgtaaaa acccgcttcg
     4201 gcgggttttt ttatgggggg agtttaggga aagagcattt gtcagctgcg tctcaagcag
     4261 ttacagagat gttacgaacc cccaggacat ccgagaatgc gaggcgatgg agggtacaac
     4321 ccgagcggcc gcttatttgc cgactacctt ggtgatctcg cctttcacgt agtggacaaa
     4381 ttcttccaac tgatctgcgc gcgaggccaa gcgatcttct tcttgtccaa gataagcctg
     4441 tctagcttca agtatgacgg gctgatactg ggccggcagg cgctccattg cccagtcggc
     4501 agcgacatcc ttcggcgcga ttttgccggt tactgcgctg taccaaatgc gggacaacgt
     4561 aagcactaca tttcgctcat cgccagccca gtcgggcggc gagttccata gcgttaaggt
     4621 ttcatttagc gcctcaaata gatcctgttc aggaaccgga tcaaagagtt cctccgccgc
     4681 tggacctacc aaggcaacgc tatgttctct tgcttttgtc agcaagatag ccagatcaat
     4741 gtcgatcgtg gctggctcga agatacccgc aagaatgtca ttgcgctgcc attctccaaa
     4801 ttgcagttcg cgcttagctg gataacgcca cggaatgatg tcgtcgtgca caacaatggt
     4861 gacttctaca gcgcggagaa tctcgctctc tccaggggaa gccgaagttt ccaaaaggtc
     4921 gttgatcaaa gctcgccgcg ttgtttcatc aagccttacg gtcaccgtaa ccagcaaatc
     4981 aatatcactg tgtggcttca ggccgccatc cactgcggag ccgtacaaat gtacggccag
     5041 caacgtcggt tcgagatggc gctcgatgac gccaactacc tctgatagtt gagtcgatac
     5101 ttcggcgatc accgcttccc tcataatgtt taactttgtt ttagggcgac tgccctgctg
     5161 cgtaacatcg ttgctgctcc ataacatcaa acatcgaccc acggcgtaac gcgcttgctg
     5221 cttggatgcc cgaggcatag actgtacccc aaaaaaacag tcataacaag ccatgaaaac
     5281 cgccactgcg ccgttaccac cgctgcgttc ggtcaaggtt ctggaccagt tgcgtgagcg
     5341 catacgctac ttgcattaca gcttacgaac cgaacaggct tatgtccact gggttcgtgc
     5401 cttcatccgt ttccacggtg tgcgtcaccc ggcaaccttg ggtagcagcg aagtcgaggc
     5461 atttctgtcc tggctggtca tgaccaaaat cccttaacgt gagtcagcct gccgccttgg
     5521 gccgggtgat gtcgtacttg cccgccgcga actcggttac cgtccagccc agcgcgacca
     5581 gctccggcaa cgcctcgcgc acccgcttgc ggcgcttgcg catggtcgaa ccactggcct
     5641 ctgacggcca gacatagccg cacaaggtat ctatggaagc cttgccggtt ttgccggggt
     5701 cgatccagcc acacagccgc tggtgcagca ggcgggcggt ttcgctgtcc agcgcccgca
     5761 cctcgtccat gctgatgcgc acatgctggc cgccacccat gacggcctgc gcgatcaagg
     5821 ggttcagggc cacgtacagg cgcccgtccg cctcgtcgct ggcgtactcc gacagcagcc
     5881 gaaacccctg ccgcttgcgg ccattctggg cgatgatgga taccttccaa aggcgctcga
     5941 tgcagtcctg tatgtgcttg agcgccccac cactatcgac ctctgccccg atttcctttg
     6001 ccagcgcccg atagctacct ttgaccacat ggcattcagc ggtgacggcc tcccacttgg
     6061 gttccaggaa cagccggagc tgccgtccgc cttcggtctt gggttccggg ccaagcacta
     6121 ggccattagg cccagccatg gccaccagcc cttgcaggat gcgcagatca tcagcgccca
     6181 gcggctccgg gccgctgaac tcgatccgct tgccgtcgcc gtagtcatac gtcacgtcca
     6241 gcttgctgcg cttgcgctcg ccccgcttga gggcacggaa caggccgggg gccagacagt
     6301 gcgccgggtc gtgccggacg tggctgaggc tgtgcttgtt cttaggcttc accacggggc
     6361 acccccttgc tcttgcgctg cctctccagc acggcgggct tgagcacccc gccgtcatgc
     6421 cgcctgaacc accgatcagc gaacggtgcg ccatagttgg ccttgctcac accgaagcgg
     6481 acgaagaacc ggcgctggtc gtcgtccaca ccccattcct cggcctcggc gctggtcatg
     6541 ctcgacaggt aggactgcca gcggatgtta tcgaccagta ccgagctgcc ccggctggcc
     6601 tgctgctggt cgcctgcgcc catcatggcc gcgcccttgc tggcatggtg caggaacacg
     6661 atagagcacc cggtatcggc ggcgatggcc tccatgcgac cgatgacctg ggccatgggg
     6721 ccgctggcgt tttcttcctc gatgtggaac cggcgcagcg tgtccagcac catcaggcgg
     6781 cggccctcgg cggcgcgctt gaggccgtcg aaccactccg gggccatgat gttgggcagg
     6841 ctgccgatca gcggctggat cagcaggccg tcagccacgg cttgccgttc ctcggcgctg
     6901 aggtgcgccc caagggcgtg caggcggtga tgaatggcgg tgggcgggtc ttcggcgggc
     6961 aggtagatca ccgggccggt gggcagttcg cccacctcca gcagatccgg cccgcctgca
     7021 atctgtgcgg ccagttgcag ggccagcatg gatttaccgg caccaccggg cgacaccagc
     7081 gccccgaccg taccggccac catgttgggc aaaacgtagt ccagcggtgg cggcgctgct
     7141 gcgaacgcct ccagaatatt gataggctta tgggtagcca ttgattgcct cctttgcagg
     7201 cagttggtgg ttaggcgctg gcggggtcac tacccccgcc ctgcgccgct ctgagttctt
     7261 ccaggcactc gcgcagcgcc tcgtattcgt cgtcggtcag ccagaacttg cgctgacgca
     7321 tccctttggc cttcatgcgc tcggcatatc gcgcttggcg tacagcgtca gggctggcca
     7381 gcaggtcgcc ggtctgcttg tccttttggt ctttcatatc agtcaccgag aaacttgcc
//
