LOCUS       sequence-208081-        8054 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BTK assembled broad-host-range plasmid encoding GFP driven by CP32 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8054)
  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 8054)
  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: pBTK620
FEATURES             Location/Qualifiers
     source          1..8054
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      114..185
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      201..228
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(274..1089)
                     /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(274..1089)
                     /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     997..1016
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1081..1100
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(1264..2115)
                     /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(2102..2941)
                     /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(3452..4423)
                     /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            5662..5749
                     /label=RSF1010 oriT
                     /note="origin of transfer of the broad-host-range plasmid 
                     RSF1010 (Scholz et al., 1989)"
     rep_origin      6091..6485
                     /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)"
     terminator      7932..7979
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 actgcctggc gactcacaac taagcaagac agccggaacc agcgccggcg aacaccactg
       61 catatatggc atatcacaac agtccaacta gtgcactgca gtacaccagg catcaaataa
      121 aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg
      181 ctctctacta gagtcacact ggctcacctt cgggtgggcc tttctgcgtt tataccgagc
      241 ggccgcgtgt tacaaccaat taaccaattc tgattagaaa aactcatcga gcatcaaatg
      301 aaactgcaat ttattcatat caggattatc aataccatat ttttgaaaaa gccgtttctg
      361 taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct ggtatcggtc
      421 tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt caaaaataag
      481 gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg gcaaaagctt
      541 atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat caaaatcact
      601 cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcgaggcgaa atacgcgatc
      661 gctgttaaaa ggacaattac aaacaggaat cgaatgcaac cggcgcagga acactgccag
      721 cgcatcaaca atattttcac ctgaatcagg atattcttct aatacctgga atgctgtttt
      781 cccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa aatgcttgat
      841 ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg accatctcat ctgtaacatc
      901 attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg gcttcccata
      961 caatcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt tatacccata
     1021 taaatcagca tccatgttgg aatttaatcg cggcctggag caagacgttt cccgttgaat
     1081 atggctcata acaccccttg tattactgtt tatgtaagca gacagtttta ttgttcatga
     1141 tgatatattt ttatcttgtg caatgtaaca tcagagattt tgagacacaa cgtggctttg
     1201 ttgaataaat cgaacttttg ctgagttgaa ggatcagtca tgaccaaaat cccttaacgt
     1261 gagtcagcct gccgccttgg gccgggtgat gtcgtacttg cccgccgcga actcggttac
     1321 cgtccagccc agcgcgacca gctccggcaa cgcctcgcgc acccgcttgc ggcgcttgcg
     1381 catggtcgaa ccactggcct ctgacggcca gacatagccg cacaaggtat ctatggaagc
     1441 cttgccggtt ttgccggggt cgatccagcc acacagccgc tggtgcagca ggcgggcggt
     1501 ttcgctgtcc agcgcccgca cctcgtccat gctgatgcgc acatgctggc cgccacccat
     1561 gacggcctgc gcgatcaagg ggttcagggc cacgtacagg cgcccgtccg cctcgtcgct
     1621 ggcgtactcc gacagcagcc gaaacccctg ccgcttgcgg ccattctggg cgatgatgga
     1681 taccttccaa aggcgctcga tgcagtcctg tatgtgcttg agcgccccac cactatcgac
     1741 ctctgccccg atttcctttg ccagcgcccg atagctacct ttgaccacat ggcattcagc
     1801 ggtgacggcc tcccacttgg gttccaggaa cagccggagc tgccgtccgc cttcggtctt
     1861 gggttccggg ccaagcacta ggccattagg cccagccatg gccaccagcc cttgcaggat
     1921 gcgcagatca tcagcgccca gcggctccgg gccgctgaac tcgatccgct tgccgtcgcc
     1981 gtagtcatac gtcacgtcca gcttgctgcg cttgcgctcg ccccgcttga gggcacggaa
     2041 caggccgggg gccagacagt gcgccgggtc gtgccggacg tggctgaggc tgtgcttgtt
     2101 cttaggcttc accacggggc acccccttgc tcttgcgctg cctctccagc acggcgggct
     2161 tgagcacccc gccgtcatgc cgcctgaacc accgatcagc gaacggtgcg ccatagttgg
     2221 ccttgctcac accgaagcgg acgaagaacc ggcgctggtc gtcgtccaca ccccattcct
     2281 cggcctcggc gctggtcatg ctcgacaggt aggactgcca gcggatgtta tcgaccagta
     2341 ccgagctgcc ccggctggcc tgctgctggt cgcctgcgcc catcatggcc gcgcccttgc
     2401 tggcatggtg caggaacacg atagagcacc cggtatcggc ggcgatggcc tccatgcgac
     2461 cgatgacctg ggccatgggg ccgctggcgt tttcttcctc gatgtggaac cggcgcagcg
     2521 tgtccagcac catcaggcgg cggccctcgg cggcgcgctt gaggccgtcg aaccactccg
     2581 gggccatgat gttgggcagg ctgccgatca gcggctggat cagcaggccg tcagccacgg
     2641 cttgccgttc ctcggcgctg aggtgcgccc caagggcgtg caggcggtga tgaatggcgg
     2701 tgggcgggtc ttcggcgggc aggtagatca ccgggccggt gggcagttcg cccacctcca
     2761 gcagatccgg cccgcctgca atctgtgcgg ccagttgcag ggccagcatg gatttaccgg
     2821 caccaccggg cgacaccagc gccccgaccg taccggccac catgttgggc aaaacgtagt
     2881 ccagcggtgg cggcgctgct gcgaacgcct ccagaatatt gataggctta tgggtagcca
     2941 ttgattgcct cctttgcagg cagttggtgg ttaggcgctg gcggggtcac tacccccgcc
     3001 ctgcgccgct ctgagttctt ccaggcactc gcgcagcgcc tcgtattcgt cgtcggtcag
     3061 ccagaacttg cgctgacgca tccctttggc cttcatgcgc tcggcatatc gcgcttggcg
     3121 tacagcgtca gggctggcca gcaggtcgcc ggtctgcttg tccttttggt ctttcatatc
     3181 agtcaccgag aaacttgccg gggccgaaag gcttgtcttc gcggaacaag gacaaggtgc
     3241 agccgtcaag gttaaggctg gccatatcag cgactgaaaa gcggccagcc tcggccttgt
     3301 ttgacgtata accaaagcca ccgggcaacc aatagccctt gtcacttttg atcaggtaga
     3361 ccgaccctga agcgcttttt tcgtattcca taaaaccccc ttctgtgcgt gagtactcat
     3421 agtataacag gcgtgagtac caacgcaagc actacatgct gaaatctggc ccgcccctgt
     3481 ccatgcctcg ctggcggggt gccggtgccc gtgccagctc ggcccgcgca agctggacgc
     3541 tgggcagacc catgaccttg ctgacggtgc gctcgatgta atccgcttcg tggccgggct
     3601 tgcgctctgc cagcgctggg ctggcctcgg ccatggcctt gccgatttcc tcggcactgc
     3661 ggccccggct ggccagcttc tgcgcggcga taaagtcgca cttgctgagg tcatgaccga
     3721 agcgcttgac cagcccggcc atctcgctgc ggtactcgtc cagcgccgtg cgccggtggc
     3781 ggctaagctg ccgctcgggc agttcgaggc tggccagcct gcgggccttc tcctgctgcc
     3841 gctgggcctg ctcgatctgc tggccagcct gctgcaccag cgccgggcca gcggtggcgg
     3901 tcttgccctt ggattcacgc agcagcaccc acggctgata accggcgcgg gtggtgtgct
     3961 tgtccttgcg gttggtgaag cccgccaagc ggccatagtg gcggctgtcg gcgctggccg
     4021 ggtcggcgtc gtactcgctg gccagcgtcc gggcaatctg cccccgaagt tcaccgcctg
     4081 cggcgtcggc caccttgacc catgcctgat agttcttcgg gctggtttcc actaccaggg
     4141 caggctcccg gccctcggct ttcatgtcat ccaggtcaaa ctcgctgagg tcgtccacca
     4201 gcaccagacc atgccgctcc tgctcggcgg gcctgatata cacgtcattg ccctgggcat
     4261 tcatccgctt gagccatggc gtgttctgga gcacttcggc ggctgaccat tcccggttca
     4321 tcatctggcc ggtgggtgcg tccctgacgc cgatatcgaa gcgctcacag cccatggcct
     4381 tgagctgtcg gcctatggcc tgcaaagtcc tgtcgttctt catcgggcca ccaagcgcag
     4441 ccagatcgag ccgtcctcgg ttgtcagtgg cgtcaggtcg agcaagagca acgatgcgat
     4501 cagcagcacc accgtaggca tcatggaagc cagcatcacg gttagccata gcttccagtg
     4561 ccacccccgc gacgcgctcc gggcgctctg cgcggcgctg ctcacctcgg cggctacctc
     4621 ccgcaactct ttggccagct ccacccatgc cgcccctgtc tggcgctggg ctttcagcca
     4681 ctccgccgcc tgcgcctcgc tggcctgctt ggtctggctc atgacctgcc gggcttcgtc
     4741 ggccagtgtc gccatgctct gggccagcgg ttcgatctgc tccgctaact cgttgatgcc
     4801 tctggatttc ttcactctgt cgattgcgtt catggtctat tgcctcccgg tattcctgta
     4861 agtcgatgat ctgggcgttg gcggtgtcga tgttcagggc cacgtctgcc cggtcggtgc
     4921 ggatgccccg gccttccatc tccaccacgt tcggccccag gtgaacaccg ggcaggcgct
     4981 cgatgccctg cgcctcaagt gttctgtggt caatgcgggc gtcgtggcca gcccgctcta
     5041 atgcccggtt ggcatggtcg gcccatgcct cgcgggtctg ctcaagccat gccttgggct
     5101 tgagcgcttc ggtcttctgt gccccgccct tctccggggt cttgccgttg taccgcttga
     5161 accactgagc ggcgggccgc tcgatgccgt cattgatccg ctcggagatc atcaggtggc
     5221 agtgcgggtt ctcgccgcca ccggcatgga tggccagcgt atacggcagg cgctcggcac
     5281 cggtcaggtg ctgggcgaac tcggacgcca gcgccttctg ctggtcgagg gtcagctcga
     5341 ccggcagggc aaattcgacc tccttgaaca gccgcccatt ggcgcgttca tacaggtcgg
     5401 cagcatccca gtagtcggcg ggccgctcga cgaactccgg catgtgcccg gattcggcgt
     5461 gcaagacttc atccatgtcg cgggcatact tgccttcgcg ctggatgtag tcggccttgg
     5521 ccctggccga ttggccgccc gacctgctgc cggttttcgc cgtaaggtga taaatcgcca
     5581 tgctgcctcg ctgttgcttt tgcttttcgg ctccatgcaa tggccctcgg agagcgcacc
     5641 gcccgaaggg tggccgttag gccagtttct cgaagagaaa ccggtaagtg cgccctcccc
     5701 tacaaagtag ggtcgggatt gccgccgctg tgcctccatg atagcctacg agacagcaca
     5761 ttaacaatgg ggtgtcaaga tggttaaggg gagcaacaag gcggcggatc ggctggccaa
     5821 gctcgaagaa caacgagcgc gaatcaatgc cgaaattcag cgggagcggg caagggaaca
     5881 gcagcaagag cgcaagaacg aaacaaggcg caaggtgctg gtgggggcca tgattttggc
     5941 caaggtgaac agcagcgagt ggccggagga tcggctcatg gcggcaatgg atgcgtacct
     6001 tgaacgcgac cacgaccgcg ccttgttcgg tctgccgcca cgccagaagg atgagccggg
     6061 ctgaatgatc gaccgagaca ggccctgcgg ggctgcacac gcgcccccac ccttcgggta
     6121 gggggaaagg ccgctaaagc ggctaaaagc gctccagcgt atttctgcgg ggtttggtgt
     6181 ggggtttagc gggctttgcc cgcctttccc cctgccgcgc agcggtgggg cggtgtgtag
     6241 cctagcgcag cgaatagacc agctatccgg cctctggccg ggcatattgg gcaagggcag
     6301 cagcgcccca caagggcgct gataaccgcg cctagtggat tattcttaga taatcatgga
     6361 tggatttttc caacaccccg ccagcccccg cccctgctgg gtttgcaggt ttgggggcgt
     6421 gacagttatt gcaggggttc gtgacagtta ttgcaggggg gcgtgacagt tattgcaggg
     6481 gttcgtgaca gttagtacgg gagtgacggg cactggctgg caatgtctag caacggcagg
     6541 catttcggct gagggtaaaa gaactttccg ctaagcgata gactgtatgt aaacacagta
     6601 ttgcaaggac gcggaacatg cctcatgtgg cggccaggac ggccagccgg gatcgggata
     6661 ctggtcgtta ccagagccac cgacccgagc aaacccttct ctatcagatc gttgacgagt
     6721 attacccggc attcgctgcg cttatggcag agcagggaaa ggaattgccg ggctatgtgc
     6781 aacgggaatt tgaagaattt ctccaatgcg ggcggctgga gcatggcttt ctacgggttc
     6841 gctgcgagtc ttgccacgcc gagcacctgg tcgctttcag aaatcaatct aaagtatata
     6901 tgagtaaact tggtctgaca ggcccctgaa ttcgcatcta gatggtagag ccacaaacag
     6961 ccggtacaag caacgatctc caggaccatc tgaatcatgc gcggatgaca cgaactcacg
     7021 acggcgatca cagacattaa cccacagtac agacactgcg acaacgtggc aattcgtcgc
     7081 aataccgtct cactgaactg gccgataatt gcagacgaac gcatacggga gtttattctt
     7141 gacatattgc cggtgtgttg gtataataac ttagtactgt tatacagaaa cagaggagat
     7201 attacatatg agtaaaggag aagagctttt cacaggagtt gtcccaatcc tcgtggaatt
     7261 agacggtgat gttaatgggc acaagttctc tgtcagtgga gagggtgaag gcgacgcaac
     7321 atatggcaag ctgaccctta aatttatttg caccacgggt aaactacctg ttccatggcc
     7381 aacactggtc actacgttcg ggtatggggt tcagtgcttt gcgcgctacc cagatcacat
     7441 gaaacagcac gactttttca agagtgcaat gcccgaaggc tatgtacagg agagaaccat
     7501 cttttttaag gatgacggca actataagac acgcgccgaa gtgaagttcg agggtgatac
     7561 ccttgttaat agaatcgagt taaagggtat tgactttaag gaagatggaa atattttagg
     7621 ccacaaactg gaatataact ataactccca taatgtgtac attatggccg acaagcaaaa
     7681 gaacggtatc aaggttaact tcaagatcag acacaacatt gaggatggaa gcgttcaact
     7741 agccgaccat taccaacaaa acaccccaat tggcgatggg cctgtgctgt taccagacaa
     7801 ccattacctg tccactcaat ctgccctttc gaaagatccc aacgaaaagc gcgaccacat
     7861 ggtccttctt gagtttgtca cggctgctgg gattacacac ggcatggatg aactatacaa
     7921 ataaatccta actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc
     7981 tggctgagca tgagacggaa atctgctcgt cagtggtgct cacactgacg aatcatgtac
     8041 agatcatacc gatg
//
