LOCUS       sequence-223001-        8054 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BTK assembled broad-host-range plasmid encoding GFP driven by CP6 
            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: pBTK621
FEATURES             Location/Qualifiers
     source          1..8054
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      42..113
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      129..156
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(202..1017)
                     /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(202..1017)
                     /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     925..944
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1009..1028
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(1192..2043)
                     /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
                     TADAMVVKGSYRALAKEIGAEVDSGGALKHIQDCIERLWKVSIIAQNGRKRQGFRLLSE
                     YASDEADGRLYVALNPLIAQAVMGGGQHVRISMDEVRALDSETARLLHQRLCGWIDPGK
                     TGKASIDTLCGYVWPSEASGSTMRKRRQRVREALPELVALGWTVTEFAAGKYDITRPKA
                     AG"
     CDS             complement(2030..2869)
                     /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(3380..4351)
                     /codon_start=1
                     /gene="repB"
                     /product="replication protein B of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepB
                     /translation="MKNDRTLQAIGRQLKAMGCERFDIGVRDATTGQMMNREWSAAEVL
                     QNTPWLKRMNAQGNDVYIRPAEQERHGLVLVDDLSEFDLDDMKAEGREPALVVETSPKN
                     YQAWVKVADAAGGELRGQIARTLASEYDADPASADSRHYGRLAGFTNRKDKHTTRAGYQ
                     PWVLLRESKGKTATAGPALVQQAGQQIEQAQRQQEKARRLASLELPERQLSRHRRTALD
                     EYRSEMAGLVKRFGDDLSKCDFIAAQKLASRGRSAEEIGKAMAEASPALAERKPGHEAD
                     YIERTVSKVMGLPSVQLARAELARAPAPRQRGMDRGGPDFSM"
     oriT            5590..5677
                     /label=RSF1010 oriT
                     /note="origin of transfer of the broad-host-range plasmid 
                     RSF1010 (Scholz et al., 1989)"
     rep_origin      6019..6413
                     /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      7860..7907
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 atcacaacag tccaactagt gcactgcagt acaccaggca tcaaataaaa cgaaaggctc
       61 agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctctactaga
      121 gtcacactgg ctcaccttcg ggtgggcctt tctgcgttta taccgagcgg ccgcgtgtta
      181 caaccaatta accaattctg attagaaaaa ctcatcgagc atcaaatgaa actgcaattt
      241 attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga
      301 aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac
      361 tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga
      421 gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagcttat gcatttcttt
      481 ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa
      541 accgttattc attcgtgatt gcgcctgagc gaggcgaaat acgcgatcgc tgttaaaagg
      601 acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat
      661 attttcacct gaatcaggat attcttctaa tacctggaat gctgttttcc cggggatcgc
      721 agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg
      781 cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat tggcaacgct
      841 acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca atcgatagat
      901 tgtcgcacct gattgcccga cattatcgcg agcccattta tacccatata aatcagcatc
      961 catgttggaa tttaatcgcg gcctggagca agacgtttcc cgttgaatat ggctcataac
     1021 accccttgta ttactgttta tgtaagcaga cagttttatt gttcatgatg atatattttt
     1081 atcttgtgca atgtaacatc agagattttg agacacaacg tggctttgtt gaataaatcg
     1141 aacttttgct gagttgaagg atcagtcatg accaaaatcc cttaacgtga gtcagcctgc
     1201 cgccttgggc cgggtgatgt cgtacttgcc cgccgcgaac tcggttaccg tccagcccag
     1261 cgcgaccagc tccggcaacg cctcgcgcac ccgctggcgg cgcttgcgca tggtcgaacc
     1321 actggcctct gacggccaga catagccgca caaggtatct atggaagcct tgccggtttt
     1381 gccggggtcg atccagccac acagccgctg gtgcagcagg cgggcggttt cgctgtccag
     1441 cgcccgcacc tcgtccatgc tgatgcgcac atgctggccg ccacccatga cggcctgcgc
     1501 gatcaagggg ttcagggcca cgtacaggcg cccgtccgcc tcgtcgctgg cgtactccga
     1561 cagcagccga aacccctgcc gcttgcggcc attctgggcg atgatggata ccttccaaag
     1621 gcgctcgatg cagtcctgta tgtgcttgag cgccccacca ctatcgacct ctgccccgat
     1681 ttcctttgcc agcgcccgat agctaccttt gaccaccatg gcatcagcgg tgacggcctc
     1741 ccacttgggt tccaggaaca gccggagctg ccgtccgcct tcggtcttgg gttccgggcc
     1801 aagcactagg ccattaggcc cagccatggc caccagccct tgcaggatgc gcagatcatc
     1861 agcgcccagc ggctccgggc cgctgaactc gatccgcttg ccgtcgccgt agtcatacgt
     1921 cacgtccagc ttgctgcgct tgcgctcgcc ccgcttgagg gcacggaaca ggccgggggc
     1981 cagacagtgc gccgggtcgt gccggacgtg gctgaggctg tgcttgttct taggcttcac
     2041 cacggggcac ccccttgctc ttgcgctgcc tctccagcac ggcgggcttg agcaccccgc
     2101 cgtcatgccg cctgaaccac cgatcagcga acggtgcgcc atagttggcc ttgctcacac
     2161 cgaagcggac gaagaaccgg cgctggtcgt cgtccacacc ccattcctcg gcctcggcgc
     2221 tggtcatgct cgacaggtag gactgccagc ggatgttatc gaccagtacc gagctgcccc
     2281 ggctggcctg ctgctggtcg cctgcgccca tcatggccgc gcccttgctg gcatggtgca
     2341 ggaacacgat agagcacccg gtatcggcgg cgatggcctc catgcgaccg atgacctggg
     2401 ccatggggcc gctggcgttt tcttcctcga tgtggaaccg gcgcagcgtg tccagcacca
     2461 tcaggcggcg gccctcggcg gcgcgcttga ggccgtcgaa ccactccggg gccatgatgt
     2521 tgggcaggct gccgatcagc ggctggatca gcaggccgtc agccacggct tgccgttcct
     2581 cggcgctgag gtgcgcccca agggcgtgca ggcggtgatg aatggcggtg ggcgggtctt
     2641 cggcgggcag gtagatcacc gggccggtgg gcagttcgcc cacctccagc agatccggcc
     2701 cgcctgcaat ctgtgcggcc agttgcaggg ccagcatgga tttaccggca ccaccgggcg
     2761 acaccagcgc cccgaccgta ccggccacca tgttgggcaa aacgtagtcc agcggtggcg
     2821 gcgctgctgc gaacgcctcc agaatattga taggcttatg ggtagccatt gattgcctcc
     2881 tttgcaggca gttggtggtt aggcgctggc ggggtcacta cccccgccct gcgccgctct
     2941 gagttcttcc aggcactcgc gcagcgcctc gtattcgtcg tcggtcagcc agaacttgcg
     3001 ctgacgcatc cctttggcct tcatgcgctc ggcatatcgc gcttggcgta cagcgtcagg
     3061 gctggccagc aggtcgccgg tctgcttgtc cttttggtct ttcatatcag tcaccgagaa
     3121 acttgccggg gccgaaaggc ttgtcttcgc ggaacaagga caaggtgcag ccgtcaaggt
     3181 taaggctggc catatcagcg actgaaaagc ggccagcctc ggccttgttt gacgtataac
     3241 caaagccacc gggcaaccaa tagcccttgt cacttttgat caggtagacc gaccctgaag
     3301 cgcttttttc gtattccata aaaccccctt ctgtgcgtga gtactcatag tataacaggc
     3361 gtgagtacca acgcaagcac tacatgctga aatctggccc gcccctgtcc atgcctcgct
     3421 ggcggggtgc cggtgcccgt gccagctcgg cccgcgcaag ctggacgctg ggcagaccca
     3481 tgaccttgct gacggtgcgc tcgatgtaat ccgcttcgtg gccgggcttg cgctctgcca
     3541 gcgctgggct ggcctcggcc atggccttgc cgatttcctc ggcactgcgg ccccggctgg
     3601 ccagcttctg cgcggcgata aagtcgcact tgctgaggtc atcaccgaag cgcttgacca
     3661 gcccggccat ctcgctgcgg tactcgtcca gcgccgtgcg ccggtggcgg ctaagctgcc
     3721 gctcgggcag ttcgaggctg gccagcctgc gggccttctc ctgctgccgc tgggcctgct
     3781 cgatctgctg gccagcctgc tgcaccagcg ccgggccagc ggtggcggtc ttgcccttgg
     3841 attcacgcag cagcacccac ggctgataac cggcgcgggt ggtgtgcttg tccttgcggt
     3901 tggtgaagcc cgccaagcgg ccatagtggc ggctgtcggc gctggccggg tcggcgtcgt
     3961 actcgctggc cagcgtccgg gcaatctgcc cccgaagttc accgcctgcg gcgtcggcca
     4021 ccttgaccca tgcctgatag ttcttcgggc tggtttccac taccagggca ggctcccggc
     4081 cctcggcttt catgtcatcc aggtcaaact cgctgaggtc gtccaccagc accagaccat
     4141 gccgctcctg ctcggcgggc ctgatataca cgtcattgcc ctgggcattc atccgcttga
     4201 gccatggcgt gttctggagc acttcggcgg ctgaccattc ccggttcatc atctggccgg
     4261 tggtggcgtc cctgacgccg atatcgaagc gctcacagcc catggccttg agctgtcggc
     4321 ctatggcctg caaagtcctg tcgttcttca tcgggccacc aagcgcagcc agatcgagcc
     4381 gtcctcggtt gtcagtggcg tcaggtcgag caagagcaac gatgcgatca gcagcaccac
     4441 cgtaggcatc atggaagcca gcatcacggt tagccatagc ttccagtgcc acccccgcga
     4501 cgcgctccgg gcgctctgcg cggcgctgct cacctcggcg gctacctccc gcaactcttt
     4561 ggccagctcc acccatgccg cccctgtctg gcgctgggct ttcagccact ccgccgcctg
     4621 cgcctcgctg gcctgctggg tctggctcat gacctgccgg gcttcgtcgg ccagtgtcgc
     4681 catgctctgg gccagcggtt cgatctgctc cgctaactcg ttgatgcctc tggatttctt
     4741 cactctgtcg attgcgttca tggtctattg cctcccggta ttcctgtaag tcgatgatct
     4801 gggcgttggc ggtgtcgatg ttcagggcca cgtctgcccg gtcggtgcgg atgccccggc
     4861 cttccatctc caccacgttc ggccccaggt gaacaccggg caggcgctcg atgccctgcg
     4921 cctcaagtgt tctgtggtca atgcgggcgt cgtggccagc ccgctctaat gcccggttgg
     4981 catggtcggc ccatgcctcg cgggtctgct caagccatgc cttgggcttg agcgcttcgg
     5041 tcttctgtgc cccgcccttc tccggggtct tgccgttgta ccgcttgaac cactgagcgg
     5101 cgggccgctc gatgccgtca ttgatccgct cggagatcat caggtggcag tgcgggttct
     5161 cgccgccacc ggcatggatg gccagcgtat acggcaggcg ctcggcaccg gtcaggtgct
     5221 gggcgaactc ggacgccagc gccttctgct ggtcgagggt cagctcgacc ggcagggcaa
     5281 attcgacctc cttgaacagc cgcccattgg cgcgttcata caggtcggca gcatcccagt
     5341 agtcggcggg ccgctcgacg aactccggca tgtgcccgga ttcggcgtgc aagacttcat
     5401 ccatgtcgcg ggcatacttg ccttcgcgct ggatgtagtc ggccttggcc ctggccgatt
     5461 ggccgcccga cctgctgccg gttttcgccg taaggtgata aatcgccatg ctgcctcgct
     5521 gttgcttttg cttttcggct ccatgcaatg gccctcggag agcgcaccgc ccgaagggtg
     5581 gccgttaggc cagtttctcg aagagaaacc ggtaagtgcg ccctccccta caaagtaggg
     5641 tcgggattgc cgccgctgtg cctccatgat agcctacgag acagcacatt aacaatgggg
     5701 tgtcaagatg gttaagggga gcaacaaggc ggcggatcgg ctggccaagc tcgaagaaca
     5761 acgagcgcga atcaatgccg aaattcagcg ggtgcgggca agggaacagc agcaagagcg
     5821 caagaacgaa acaaggcgca aggtgctggt gggggccatg attttggcca aggtgaacag
     5881 cagcgagtgg ccggaggatc ggctcatggc ggcaatggat gcgtaccttg aacgcgacca
     5941 cgaccgcgcc ttgttcggtc tgccgccacg ccagaaggat gagccgggct gaatgatcga
     6001 ccgagacagg ccctgcgggg ctgcacacgc gcccccaccc ttcgggtagg gggaaaggcc
     6061 gctaaagcgg ctaaaagcgc tccagcgtat ttctgcgggg tttggtgtgg ggtttagcgg
     6121 gctttgcccg cctttccccc tgccgcgcag cggtggggcg gtgtgtagcc tagcgcagcg
     6181 aatagaccag ctatccggcc tctggccggg catattgggc aagggcagca gcgccccaca
     6241 agggcgctga taaccgcgcc tagtggatta ttcttagata atcatggatg gatttttcca
     6301 acaccccgcc agcccccgcc cctgctgggt ttgcaggttt gggggcgtga cagttattgc
     6361 aggggttcgt gacagttatt gcaggggggc gtgacagtta ttgcaggggt tcgtgacagt
     6421 tagtacggga gtgacgggca ctggctggca atgtctagca acggcaggca tttcggctga
     6481 gggtaaaaga actttccgct aagcgataga ctgtatgtaa acacagtatt gcaaggacgc
     6541 ggaacatgcc tcatgtggcg gccaggacgg ccagccggga tcgggatact ggtcgttacc
     6601 agagccaccg acccgagcaa acccttctct atcagatcgt tgacgagtat tacccggcat
     6661 tcgctgcgct tatggcagag cagggaaagg aattgccggg ctatgtgcaa cgggaatttg
     6721 aagaatttct ccaatgcggg cggctggagc atggctttct acgggttcgc tgcgagtctt
     6781 gccacgccga gcacctggtc gctttcagaa atcaatctaa agtatatatg agtaaacttg
     6841 gtctgacagg cccctgaatt cgcatctaga tggtagagcc acaaacagcc ggtacaagca
     6901 acgatctcca ggaccatctg aatcatgcgc ggatgacacg aactcacgac ggcgatcaca
     6961 gacattaacc cacagtacag acactgcgac aacgtggcaa ttcgtcgcaa taccgtctca
     7021 ctgaactggc cgataattgc agacgaacgc atgtgggagt ttattcttga cacagatatt
     7081 tccggatgat ataataactg agtactgtta tacagaaaca gaggagatat tacatatgag
     7141 taaaggagaa gagcttttca caggagttgt cccaatcctc gtggaattag acggtgatgt
     7201 taatgggcac aagttctctg tcagtggaga gggtgaaggc gacgcaacat atggcaagct
     7261 gacccttaaa tttatttgca ccacgggtaa actacctgtt ccatggccaa cactggtcac
     7321 tacgttcggg tatggggttc agtgctttgc gcgctaccca gatcacatga aacagcacga
     7381 ctttttcaag agtgcaatgc ccgaaggcta tgtacaggag agaaccatct tttttaagga
     7441 tgacggcaac tataagacac gcgccgaagt gaagttcgag ggtgataccc ttgttaatag
     7501 aatcgagtta aagggtattg actttaagga agatggaaat attttaggcc acaaactgga
     7561 atataactat aactcccata atgtgtacat tatggccgac aagcaaaaga acggtatcaa
     7621 ggttaacttc aagatcagac acaacattga ggatggaagc gttcaactag ccgaccatta
     7681 ccaacaaaac accccaattg gcgatgggcc tgtgctgtta ccagacaacc attacctgtc
     7741 cactcaatct gccctttcga aagatcccaa cgaaaagcgc gaccacatgg tccttcttga
     7801 gtttgtcacg gctgctggga ttacacacgg catggatgaa ctatacaaat aaatcctaac
     7861 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg gctgagcatg
     7921 agacggaaat ctgctcgtca gtggtgctca cactgacgaa tcatgtacag atcataccga
     7981 tgactgcctg gcgactcaca actaagcaag acagccggaa ccagcgccgg cgaacaccac
     8041 tgcatatatg gcat
//
