LOCUS       sequence-218136-        8802 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BTK assembled broad-host-range plasmid encoding GFP driven by PA2 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8802)
  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 8802)
  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: pBTK509
FEATURES             Location/Qualifiers
     source          1..8802
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(158..1009)
                     /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
                     GDGKRIEFSGPEPLGADDLRILQGLVAMAGPNGLVLGPEPKTEGGRQLRLFLEPKREAV
                     TADAMVVKGSYRALAKEIGAEVDSGGALKHIQDCIERLWKVSIIAQNGRKRQGFRLLSE
                     YASDEADGRLYVALNPLIAQAVMGGGQHVRISMDEVRALDSETARLLHQRLCGWIDPGK
                     TGKASIDTLCGYVWPSEASGSTMRKRRQRVREALPELVALGWTVTEFAAGKYDITRPKA
                     AG"
     CDS             complement(996..1835)
                     /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(2346..3317)
                     /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            4556..4643
                     /label=RSF1010 oriT
                     /note="origin of transfer of the broad-host-range plasmid 
                     RSF1010 (Scholz et al., 1989)"
     rep_origin      4985..5379
                     /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      complement(7094..7140)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(7149..7772)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     7680..7699
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(7812..7831)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(7813..7831)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             complement(join(7942..8733,8734..8802))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     8565..8584
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct gtcatgacca aaatccctta acgtgagtca gcctgccgcc ttgggccggg
      181 tgatgtcgta cttgcccgcc gcgaactcgg ttaccgtcca gcccagcgcg accagctccg
      241 gcaacgcctc gcgcacccgc tggcggcgct tgcgcatggt cgaaccactg gcctctgacg
      301 gccagacata gccgcacaag gtatctatgg aagccttgcc ggttttgccg gggtcgatcc
      361 agccacacag ccgctggtgc agcaggcggg cggtttcgct gtccagcgcc cgcacctcgt
      421 ccatgctgat gcgcacatgc tggccgccac ccatgacggc ctgcgcgatc aaggggttca
      481 gggccacgta caggcgcccg tccgcctcgt cgctggcgta ctccgacagc agccgaaacc
      541 cctgccgctt gcggccattc tgggcgatga tggatacctt ccaaaggcgc tcgatgcagt
      601 cctgtatgtg cttgagcgcc ccaccactat cgacctctgc cccgatttcc tttgccagcg
      661 cccgatagct acctttgacc accatggcat cagcggtgac ggcctccctc ttgggttcca
      721 ggaacagccg gagctgccgt ccgccttcgg tcttgggttc cgggccaagc actaggccat
      781 taggcccagc catggccacc agcccttgca ggatgcgcag atcatcagcg cccagcggct
      841 ccgggccgct gaactcgatc cgcttgccgt cgccgtagtc atacgtcacg tccagcttgc
      901 tgcgcttgcg ctcgccccgc ttgagggcac ggaacaggcc gggggccaga cagtgcgccg
      961 ggtcgtgccg gacgtggctg aggctgtgct tgttcttagg cttcaccacg gggcaccccc
     1021 ttgctcttgc gctgcctctc cagcacggcg ggcttgagca ccccgccgtc atgccgcctg
     1081 aaccaccgat cagcgaacgg tgcgccatag ttggccttgc tcacaccgaa gcggacgaag
     1141 aaccggcgct ggtcgtcgtc cacaccccat tcctcggcct cggcgctggt catgctcgac
     1201 aggtaggact gccagcggat gttatcgacc agtaccgagc tgccccggct ggcctgctgc
     1261 tggtcgcctg cgcccatcat ggccgcgccc ttgctggcat ggtgcaggaa cacgatagag
     1321 cacccggtat cggcggcgat ggcctccatg cgaccgatga cctgggccat ggggccgctg
     1381 gcgttttctt cctcgatgtg gaaccggcgc agcgtgtcca gcaccatcag gcggcggccc
     1441 tcggcggcgc gcttgaggcc gtcgaaccac tccggggcca tgatgttggg caggctgccg
     1501 atcagcggct ggatcagcag gccgtcagcc acggcttgcc gttcctcggc gctgaggtgc
     1561 gccccaaggg cgtgcaggcg gtgatgaatg gcggtgggcg ggtcttcggc gggcaggtag
     1621 atcaccgggc cggtgggcag ttcgcccacc tccagcagat ccggcccgcc tgcaatctgt
     1681 gcggccagtt gcagggccag catggattta ccggcaccac cgggcgacac cagcgccccg
     1741 accgtaccgg ccaccatgtt gggcaaaacg tagtccagcg gtggcggcgc tgctgcgaac
     1801 gcctccagaa tattgatagg cttatgggta gccattgatt gcctcctttg caggcagttg
     1861 gtggttaggc gctggcgggg tcactacccc cgccctgcgc cgctctgagt tcttccaggc
     1921 actcgcgcag cgcctcgtat tcgtcgtcgg tcagccagaa cttgcgctga cgcatccctt
     1981 tggccttcat gcgctcggca tatcgcgctt ggcgtacagc gtcagggctg gccagcaggt
     2041 cgccggtctg cttgtccttt tggtctttca tatcagtcac cgagaaactt gccggggccg
     2101 aaaggcttgt cttcgcggaa caaggacaag gtgcagccgt caaggttaag gctggccata
     2161 tcagcgactg aaaagcggcc agcctcggcc ttgtttgacg tataaccaaa gccaccgggc
     2221 aaccaatagc ccttgtcact tttgatcagg tagaccgacc ctgaagcgct tttttcgtat
     2281 tccataaaac ccccttctgt gcgtgagtac tcatagtata acaggcgtga gtaccaacgc
     2341 aagcactaca tgctgaaatc tggcccgccc ctgtccatgc ctcgctggcg gggtgccggt
     2401 gcccgtgcca gctcggcccg cgcaagctgg acgctgggca gacccatgac cttgctgacg
     2461 gtgcgctcga tgtaatccgc ttcgtggccg ggcttgcgct ctgccagcgc tgggctggcc
     2521 tcggccatgg ccttgccgat ttcctcggca ctgcggcccc ggctggccag cttctgcgcg
     2581 gcgataaagt cgcacttgct gaggtcatca ccgaagcgct tgaccagccc ggccatctcg
     2641 ctgcggtact cgtccagcgc cgtgcgccgg tggcggctaa gctgccgctc gggcagttcg
     2701 aggctggcca gcctgcgggc cttctcctgc tgccgctggg cctgctcgat ctgctggcca
     2761 gcctgctgca ccagcgccgg gccagcggtg gcggtcttgc ccttggattc acgcagcagc
     2821 acccacggct gataaccggc gcgggtggtg tgcttgtcct tgcggttggt gaagcccgcc
     2881 aagcggccat agtggcggct gtcggcgctg gccgggtcgg cgtcgtactc gctggccagc
     2941 gtccgggcaa tctgcccccg aagttcaccg cctgcggcgt cggccacctt gacccatgcc
     3001 tgatagttct tcgggctggt ttccactacc agggcaggct cccggccctc ggctttcatg
     3061 tcatccaggt caaactcgct gaggtcgtcc accagcacca gaccatgccg ctcctgctcg
     3121 gcgggcctga tatacacgtc attgccctgg gcattcatcc gcttgagcca tggcgtgttc
     3181 tggagcactt cggcggctga ccattcccgg ttcatcatct ggccggtggt ggcgtccctg
     3241 acgccgatat cgaagcgctc acagcccatg gccttgagct gtcggcctat ggcctgcaaa
     3301 gtcctgtcgt tcttcatcgg gccaccaagc gcagccagat cgagccgtcc tcggttgtca
     3361 gtggcgtcag gtcgagcaag agcaacgatg cgatcagcag caccaccgta ggcatcatgg
     3421 aagccagcat cacggttagc catagcttcc agtgccaccc ccgcgacgcg ctccgggcgc
     3481 tctgcgcggc gctgctcacc tcggcggcta cctcccgcaa ctctttggcc agctccaccc
     3541 atgccgcccc tgtctggcgc tgggctttca gccactccgc cgcctgcgcc tcgctggcct
     3601 gctgggtctg gctcatgacc tgccgggctt cgtcggccag tgtcgccatg ctctgggcca
     3661 gcggttcgat ctgctccgct aactcgttga tgcctctgga tttcttcact ctgtcgattg
     3721 cgttcatggt ctattgcctc ccggtattcc tgtaagtcga tgatctgggc gttggcggtg
     3781 tcgatgttca gggccacgtc tgcccggtcg gtgcggatgc cccggccttc catctccacc
     3841 acgttcggcc ccaggtgaac accgggcagg cgctcgatgc cctgcgcctc aagtgttctg
     3901 tggtcaatgc gggcgtcgtg gccagcccgc tctaatgccc ggttggcatg gtcggcccat
     3961 gcctcgcggg tctgctcaag ccatgccttg ggcttgagcg cttcggtctt ctgtgccccg
     4021 cccttctccg gggtcttgcc gttgtaccgc ttgaaccact gagcggcggg ccgctcgatg
     4081 ccgtcattga tccgctcgga gatcatcagg tggcagtgcg ggttctcgcc gccaccggca
     4141 tggatggcca gcgtatacgg caggcgctcg gcaccggtca ggtgctgggc gaactcggac
     4201 gccagcgcct tctgctggtc gagggtcagc tcgaccggca gggcaaattc gacctccttg
     4261 aacagccgcc cattggcgcg ttcatacagg tcggcagcat cccagtagtc ggcgggccgc
     4321 tcgacgaact ccggcatgtg cccggattcg gcgtgcaaga cttcatccat gtcgcgggca
     4381 tacttgcctt cgcgctggat gtagtcggcc ttggccctgg ccgattggcc gcccgacctg
     4441 ctgccggttt tcgccgtaag gtgataaatc gccatgctgc ctcgctgttg cttttgcttt
     4501 tcggctccat gcaatggccc tcggagagcg caccgcccga agggtggccg ttaggccagt
     4561 ttctcgaaga gaaaccggta agtgcgccct cccctacaaa gtagggtcgg gattgccgcc
     4621 gctgtgcctc catgatagcc tacgagacag cacattaaca atggggtgtc aagatggtta
     4681 aggggagcaa caaggcggcg gatcggctgg ccaagctcga agaacaacga gcgcgaatca
     4741 atgccgaaat tcagcgggtg cgggcaaggg aacagcagca agagcgcaag aacgaaacaa
     4801 ggcgcaaggt gctggtgggg gccatgattt tggccaaggt gaacagcagc gagtggccgg
     4861 aggatcggct catggcggca atggatgcgt accttgaacg cgaccacgac cgcgccttgt
     4921 tcggtctgcc gccacgccag aaggatgagc cgggctgaat gatcgaccga gacaggccct
     4981 gcggggctgc acacgcgccc ccacccttcg ggtaggggga aaggccgcta aagcggctaa
     5041 aagcgctcca gcgtatttct gcggggtttg gtgtggggtt tagcgggctt tgcccgcctt
     5101 tccccctgcc gcgcagcggt ggggcggtgt gtagcctagc gcagcgaata gaccagctat
     5161 ccggcctctg gccgggcata ttgggcaagg gcagcagcgc cccacaaggg cgctgataac
     5221 cgcgcctagt ggattattct tagataatca tggatggatt tttccaacac cccgccagcc
     5281 cccgcccctg ctgggtttgc aggtttgggg gcgtgacagt tattgcaggg gttcgtgaca
     5341 gttattgcag gggggcgtga cagttattgc aggggttcgt gacagttagt acgggagtga
     5401 cgggcactgg ctggcaatgt ctagcaacgg caggcatttc ggctgagggt aaaagaactt
     5461 tccgctaagc gatagactgt atgtaaacac agtattgcaa ggacgcggaa catgcctcat
     5521 gtggcggcca ggacggccag ccgggatcgg gatactggtc gttaccagag ccaccgaccc
     5581 gagcaaaccc ttctctatca gatcgttgac gagtattacc cggcattcgc tgcgcttatg
     5641 gcagagcagg gaaaggaatt gccgggctat gtgcaacggg aatttgaaga atttctccaa
     5701 tgcgggcggc tggagcatgg ctttctacgg gttcgctgcg agtcttgcca cgccgagcac
     5761 ctggtcgctt tcagaaatca atctaaagta tatatgagta aacttggtct gacaggcccc
     5821 tgaattcgca tctagactga tgagacgtgg tagagccaca aacagccggt acaagcaacg
     5881 atctccagga ccatctgaat catgcgcgga tgacacgaac tcacgacggc gatcacagac
     5941 attaacccac agtacagaca ctgcgacaac gtggcaattc gtcgcaatac aacgcacgaa
     6001 aaacaggtat tgacaacatg aagtaacatg cagtaagata caaatcgcta ggtaacacta
     6061 gcagcataca gaaacagagg agatattaca tatgagtaaa ggagaagagc ttttcacagg
     6121 agttgtccca atcctcgtgg aattagacgg tgatgttaat gggcacaagt tctctgtcag
     6181 tggagagggt gaaggcgacg caacatatgg caagctgacc cttaaattta tttgcaccac
     6241 gggtaaacta cctgttccat ggccaacact ggtcactacg ttcgggtatg gggttcagtg
     6301 ctttgcgcgc tacccagatc acatgaaaca gcacgacttt ttcaagagtg caatgcccga
     6361 aggctatgta caggagagaa ccatcttttt taaggatgac ggcaactata agacacgcgc
     6421 cgaagtgaag ttcgagggtg atacccttgt taatagaatc gagttaaagg gtattgactt
     6481 taaggaagat ggaaatattt taggccacaa actggaatat aactataact cccataatgt
     6541 gtacattatg gccgacaagc aaaagaacgg tatcaaggtt aacttcaaga tcagacacaa
     6601 cattgaggat ggaagcgttc aactagccga ccattaccaa caaaacaccc caattggcga
     6661 tgggcctgtg ctgttaccag acaaccatta cctgtccact caatctgccc tttcgaaaga
     6721 tcccaacgaa aagcgcgacc acatggtcct tcttgagttt gtcacggctg ctgggattac
     6781 acacggcatg gatgaactat acaaataaat ccgtaatcgt taatccgcaa ataacgtaaa
     6841 aacccgcttc ggcgggtttt tttatggggg gagtttaggg aaagagcatt tgtcagctgg
     6901 aaatctgctc gtcagtggtg ctcacactga cgaatcatgt acagatcata ccgatgactg
     6961 cctggcgact cacaactaag caagacagcc ggaaccagcg ccggcgaaca ccactgcata
     7021 tatggcatat cacaacagtc cacgtctcaa gcagttacag agatgttacg aaccactagt
     7081 gcactgcagt acacaaaaaa cccctcaaga cccgtttaga ggccccaagg ggttatgcta
     7141 atgctgcttt aagacccact ttcacattta agttgttttt ctaatccgca tatgatcaat
     7201 tcaaggccga ataagaaggc tggctctgca ccttggtgat caaataattc gatagcttgt
     7261 cgtaataatg gcggcatact atcagtagta ggtgtttccc tttcttcttt agcgacttga
     7321 tgctcttgat cttccaatac gcaacctaaa gtaaaatgcc ccactgcgct gagtgcatat
     7381 aatgcattct ctagtgaaaa accttgttgg cataaaaagg ctaattgatt ttcgagagtt
     7441 tcatactgtt tttctgtagg ccgtgtacct aaatgtactt ttgctccatc gcgatgactt
     7501 agtaaagcac atctaaaact tttagcgtta ttacgtaaaa aatcttgcca gctttcccct
     7561 tctaaagggc aaaagtgagt atggtgccta tctaacatct caatggctaa ggcgtcgagc
     7621 aaagcccgct tattttttac atgccaatac aatgtaggct gctctacacc tagcttctgg
     7681 gcgagtttac gggttgttaa accttcgatt ccgacctcat taagcagctc taatgcgctg
     7741 ttaatcactt tacttttatc taaacgagac atgagtatat ctccttctac aaaattattt
     7801 ctagatctct cccctatagt gagtcgtatt aatttccgag cggccgcgat tatcaaaaag
     7861 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     7921 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     7981 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     8041 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgggacccac gctcaccggc
     8101 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     8161 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     8221 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     8281 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     8341 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     8401 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     8461 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     8521 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     8581 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     8641 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     8701 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     8761 aaaaaaggga ataagggcga cacggaaatg ttgaatactc at
//
