LOCUS       sequence-218126-        7608 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BTK Type 8 Bacterial origin (broad-host-range) for golden gate 
            assembly.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7608)
  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 7608)
  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: pBTK402
FEATURES             Location/Qualifiers
     source          1..7608
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    116..134
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    141..159
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             177..188
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             195..872
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      890..961
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      977..1004
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(1061..1876)
                     /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(1061..1876)
                     /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     1784..1803
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1868..1887
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(2051..2902)
                     /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(2889..3728)
                     /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(4239..5210)
                     /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            6449..6536
                     /label=RSF1010 oriT
                     /note="origin of transfer of the broad-host-range plasmid 
                     RSF1010 (Scholz et al., 1989)"
     rep_origin      6878..7272
                     /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)"
ORIGIN
        1 catggctttc tacgggttcg ctgcgagtct tgccacgccg agcacctggt cgctttcaga
       61 aatcaatcta aagtatatat gagtaaactt ggtctgacag gccccttgag accagtccct
      121 atcagtgata gagattgaca tccctatcag tgatagagat actgagcacg gatctgaaag
      181 aggagaaagg atctatggcg agtagcgaag acgttatcaa agagttcatg cgtttcaaag
      241 ttcgtatgga aggttccgtt aacggtcacg agttcgaaat cgaaggtgaa ggtgaaggtc
      301 gtccgtacga aggtactcag accgctaaac tgaaagttac caaaggtggt ccgctgccgt
      361 tcgcttggga catcctgtcc ccgcagttcc agtacggttc caaagcttac gttaaacacc
      421 cggctgacat cccggactac ctgaaactgt ccttcccgga aggtttcaaa tgggaacgtg
      481 ttatgaactt cgaagacggt ggtgttgtta ccgttaccca ggactcctcc ctgcaagacg
      541 gtgagttcat ctacaaagtt aaactgcgtg gtactaactt cccgtccgac ggtccggtta
      601 tgcagaaaaa aaccatgggt tgggaagctt ccaccgaacg tatgtacccg gaagacggtg
      661 ctctgaaagg tgaaatcaaa atgcgtctga aactgaaaga cggtggtcac tacgacgctg
      721 aagttaaaac cacctacatg gctaaaaaac cggttcagct gccgggtgct tacaaaaccg
      781 acatcaaact ggacatcacc tcccacaacg aagactacac catcgttgaa cagtacgaac
      841 gtgctgaagg tcgtcactcc accggtgctt aataaggatc tccaggcatc aaataaaacg
      901 aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct
      961 ctactagagt cacactggct caccttcggg tgggcctttc tgcgtttata agtcggtctc
     1021 accgagcggc cgcgtgttac aaccaattaa ccaattctga ttagaaaaac tcatcgagca
     1081 tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc
     1141 gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt
     1201 atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa
     1261 aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca
     1321 aaagcttatg catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa
     1381 aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg aggcgaaata
     1441 cgcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca
     1501 ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg
     1561 ctgttttccc ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat
     1621 gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg
     1681 taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct
     1741 tcccatacaa tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat
     1801 acccatataa atcagcatcc atgttggaat ttaatcgcgg cctggagcaa gacgtttccc
     1861 gttgaatatg gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg
     1921 ttcatgatga tatattttta tcttgtgcaa tgtaacatca gagattttga gacacaacgt
     1981 ggctttgttg aataaatcga acttttgctg agttgaagga tcagtcatga ccaaaatccc
     2041 ttaacgtgag tcagcctgcc gccttgggcc gggtgatgtc gtacttgccc gccgcgaact
     2101 cggttaccgt ccagcccagc gcgaccagct ccggcaacgc ctcgcgcacc cgctggcggc
     2161 gcttgcgcat ggtcgaacca ctggcctctg acggccagac atagccgcac aaggtatcta
     2221 tggaagcctt gccggttttg ccggggtcga tccagccaca cagccgctgg tgcagcaggc
     2281 gggcggtttc gctgtccagc gcccgcacct cgtccatgct gatgcgcaca tgctggccgc
     2341 cacccatgac ggcctgcgcg atcaaggggt tcagggccac gtacaggcgc ccgtccgcct
     2401 cgtcgctggc gtactccgac agcagccgaa acccctgccg cttgcggcca ttctgggcga
     2461 tgatggatac cttccaaagg cgctcgatgc agtcctgtat gtgcttgagc gccccaccac
     2521 tatcgacctc tgccccgatt tcctttgcca gcgcccgata gctacctttg accaccatgg
     2581 catcagcggt gacggcctcc cacttgggtt ccaggaacag ccggagctgc cgtccgcctt
     2641 cggtcttggg ttccgggcca agcactaggc cattaggccc agccatggcc accagccctt
     2701 gcaggatgcg cagatcatca gcgcccagcg gctccgggcc gctgaactcg atccgcttgc
     2761 cgtcgccgta gtcatacgtc acgtccagct tgctgcgctt gcgctcgccc cgcttgaggg
     2821 cacggaacag gccgggggcc agacagtgcg ccgggtcgtg ccggacgtgg ctgaggctgt
     2881 gcttgttctt aggcttcacc acggggcacc cccttgctct tgcgctgcct ctccagcacg
     2941 gcgggcttga gcaccccgcc gtcatgccgc ctgaaccacc gatcagcgaa cggtgcgcca
     3001 tagttggcct tgctcacacc gaagcggacg aagaaccggc gctggtcgtc gtccacaccc
     3061 cattcctcgg cctcggcgct ggtcatgctc gacaggtagg actgccagcg gatgttatcg
     3121 accagtaccg agctgccccg gctggcctgc tgctggtcgc ctgcgcccat catggccgcg
     3181 cccttgctgg catggtgcag gaacacgata gagcacccgg tatcggcggc gatggcctcc
     3241 atgcgaccga tgacctgggc catggggccg ctggcgtttt cttcctcgat gtggaaccgg
     3301 cgcagcgtgt ccagcaccat caggcggcgg ccctcggcgg cgcgcttgag gccgtcgaac
     3361 cactccgggg ccatgatgtt gggcaggctg ccgatcagcg gctggatcag caggccgtca
     3421 gccacggctt gccgttcctc ggcgctgagg tgcgccccaa gggcgtgcag gcggtgatga
     3481 atggcggtgg gcgggtcttc ggcgggcagg tagatcaccg ggccggtggg cagttcgccc
     3541 acctccagca gatccggccc gcctgcaatc tgtgcggcca gttgcagggc cagcatggat
     3601 ttaccggcac caccgggcga caccagcgcc ccgaccgtac cggccaccat gttgggcaaa
     3661 acgtagtcca gcggtggcgg cgctgctgcg aacgcctcca gaatattgat aggcttatgg
     3721 gtagccattg attgcctcct ttgcaggcag ttggtggtta ggcgctggcg gggtcactac
     3781 ccccgccctg cgccgctctg agttcttcca ggcactcgcg cagcgcctcg tattcgtcgt
     3841 cggtcagcca gaacttgcgc tgacgcatcc ctttggcctt catgcgctcg gcatatcgcg
     3901 cttggcgtac agcgtcaggg ctggccagca ggtcgccggt ctgcttgtcc ttttggtctt
     3961 tcatatcagt caccgagaaa cttgccgggg ccgaaaggct tgtcttcgcg gaacaaggac
     4021 aaggtgcagc cgtcaaggtt aaggctggcc atatcagcga ctgaaaagcg gccagcctcg
     4081 gccttgtttg acgtataacc aaagccaccg ggcaaccaat agcccttgtc acttttgatc
     4141 aggtagaccg accctgaagc gcttttttcg tattccataa aacccccttc tgtgcgtgag
     4201 tactcatagt ataacaggcg tgagtaccaa cgcaagcact acatgctgaa atctggcccg
     4261 cccctgtcca tgcctcgctg gcggggtgcc ggtgcccgtg ccagctcggc ccgcgcaagc
     4321 tggacgctgg gcagacccat gaccttgctg acggtgcgct cgatgtaatc cgcttcgtgg
     4381 ccgggcttgc gctctgccag cgctgggctg gcctcggcca tggccttgcc gatttcctcg
     4441 gcactgcggc cccggctggc cagcttctgc gcggcgataa agtcgcactt gctgaggtca
     4501 tcaccgaagc gcttgaccag cccggccatc tcgctgcggt actcgtccag cgccgtgcgc
     4561 cggtggcggc taagctgccg ctcgggcagt tcgaggctgg ccagcctgcg ggccttctcc
     4621 tgctgccgct gggcctgctc gatctgctgg ccagcctgct gcaccagcgc cgggccagcg
     4681 gtggcggtct tgcccttgga ttcacgcagc agcacccacg gctgataacc ggcgcgggtg
     4741 gtgtgcttgt ccttgcggtt ggtgaagccc gccaagcggc catagtggcg gctgtcggcg
     4801 ctggccgggt cggcgtcgta ctcgctggcc agcgtccggg caatctgccc ccgaagttca
     4861 ccgcctgcgg cgtcggccac cttgacccat gcctgatagt tcttcgggct ggtttccact
     4921 accagggcag gctcccggcc ctcggctttc atgtcatcca ggtcaaactc gctgaggtcg
     4981 tccaccagca ccagaccatg ccgctcctgc tcggcgggcc tgatatacac gtcattgccc
     5041 tgggcattca tccgcttgag ccatggcgtg ttctggagca cttcggcggc tgaccattcc
     5101 cggttcatca tctggccggt ggtggcgtcc ctgacgccga tatcgaagcg ctcacagccc
     5161 atggccttga gctgtcggcc tatggcctgc aaagtcctgt cgttcttcat cgggccacca
     5221 agcgcagcca gatcgagccg tcctcggttg tcagtggcgt caggtcgagc aagagcaacg
     5281 atgcgatcag cagcaccacc gtaggcatca tggaagccag catcacggtt agccatagct
     5341 tccagtgcca cccccgcgac gcgctccggg cgctctgcgc ggcgctgctc acctcggcgg
     5401 ctacctcccg caactctttg gccagctcca cccatgccgc ccctgtctgg cgctgggctt
     5461 tcagccactc cgccgcctgc gcctcgctgg cctgctgggt ctggctcatg acctgccggg
     5521 cttcgtcggc cagtgtcgcc atgctctggg ccagcggttc gatctgctcc gctaactcgt
     5581 tgatgcctct ggatttcttc actctgtcga ttgcgttcat ggtctattgc ctcccggtat
     5641 tcctgtaagt cgatgatctg ggcgttggcg gtgtcgatgt tcagggccac gtctgcccgg
     5701 tcggtgcgga tgccccggcc ttccatctcc accacgttcg gccccaggtg aacaccgggc
     5761 aggcgctcga tgccctgcgc ctcaagtgtt ctgtggtcaa tgcgggcgtc gtggccagcc
     5821 cgctctaatg cccggttggc atggtcggcc catgcctcgc gggtctgctc aagccatgcc
     5881 ttgggcttga gcgcttcggt cttctgtgcc ccgcccttct ccggggtctt gccgttgtac
     5941 cgcttgaacc actgagcggc gggccgctcg atgccgtcat tgatccgctc ggagatcatc
     6001 aggtggcagt gcgggttctc gccgccaccg gcatggatgg ccagcgtata cggcaggcgc
     6061 tcggcaccgg tcaggtgctg ggcgaactcg gacgccagcg ccttctgctg gtcgagggtc
     6121 agctcgaccg gcagggcaaa ttcgacctcc ttgaacagcc gcccattggc gcgttcatac
     6181 aggtcggcag catcccagta gtcggcgggc cgctcgacga actccggcat gtgcccggat
     6241 tcggcgtgca agacttcatc catgtcgcgg gcatacttgc cttcgcgctg gatgtagtcg
     6301 gccttggccc tggccgattg gccgcccgac ctgctgccgg ttttcgccgt aaggtgataa
     6361 atcgccatgc tgcctcgctg ttgcttttgc ttttcggctc catgcaatgg ccctcggaga
     6421 gcgcaccgcc cgaagggtgg ccgttaggcc agtttctcga agagaaaccg gtaagtgcgc
     6481 cctcccctac aaagtagggt cgggattgcc gccgctgtgc ctccatgata gcctacgaga
     6541 cagcacatta acaatggggt gtcaagatgg ttaaggggag caacaaggcg gcggatcggc
     6601 tggccaagct cgaagaacaa cgagcgcgaa tcaatgccga aattcagcgg gtgcgggcaa
     6661 gggaacagca gcaagagcgc aagaacgaaa caaggcgcaa ggtgctggtg ggggccatga
     6721 ttttggccaa ggtgaacagc agcgagtggc cggaggatcg gctcatggcg gcaatggatg
     6781 cgtaccttga acgcgaccac gaccgcgcct tgttcggtct gccgccacgc cagaaggatg
     6841 agccgggctg aatgatcgac cgagacaggc cctgcggggc tgcacacgcg cccccaccct
     6901 tcgggtaggg ggaaaggccg ctaaagcggc taaaagcgct ccagcgtatt tctgcggggt
     6961 ttggtgtggg gtttagcggg ctttgcccgc ctttccccct gccgcgcagc ggtggggcgg
     7021 tgtgtagcct agcgcagcga atagaccagc tatccggcct ctggccgggc atattgggca
     7081 agggcagcag cgccccacaa gggcgctgat aaccgcgcct agtggattat tcttagataa
     7141 tcatggatgg atttttccaa caccccgcca gcccccgccc ctgctgggtt tgcaggtttg
     7201 ggggcgtgac agttattgca ggggttcgtg acagttattg caggggggcg tgacagttat
     7261 tgcaggggtt cgtgacagtt agtacgggag tgacgggcac tggctggcaa tgtctagcaa
     7321 cggcaggcat ttcggctgag ggtaaaagaa ctttccgcta agcgatagac tgtatgtaaa
     7381 cacagtattg caaggacgcg gaacatgcct catgtggcgg ccaggacggc cagccgggat
     7441 cgggatactg gtcgttacca gagccaccga cccgagcaaa cccttctcta tcagatcgtt
     7501 gacgagtatt acccggcatt cgctgcgctt atggcagagc agggaaagga attgccgggc
     7561 tatgtgcaac gggaatttga agaatttctc caatgcgggc ggctggag
//
