LOCUS       sequence-329553-        8293 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Genome engineering of Burkholderia.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8293)
  TITLE     Youming Zhang Lab Plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8293)
  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"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pBBR1-Rha-ETh_tji49-kan
FEATURES             Location/Qualifiers
     source          1..8293
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(60..722)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      723..1494
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             2974..3768
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3028..3047)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3638..3657
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             complement(4067..4915)
                     /codon_start=1
                     /gene="E. coli rhaR"
                     /product="transcriptional activator of rhaR and rhaS"
                     /label=rhaR
                     /note="needed for rhamnose-dependent activation of the rhaB
                     promoter"
                     /translation="MAHQLKLLKDDFFASDQQAVAVADRYPQDVFAEHTHDFCELVIVW
                     RGNGLHVLNDRPYRITRGDLFYIHADDKHSYASVNDLVLQNIIYCPERLKLNLDWQGAI
                     PGFNASAGQPHWRLGSMGMAQARQVIGQLEHESSQHVPFANEMAELLFGQLVMLLNRHR
                     YTSDSLPPTSSETLLDKLITRLAASLKSPFALDKFCDEASCSERVLRQQFRQQTGMTIN
                     QYLRQVRVCHAQYLLQHSRLLISDISTECGFEDSNYFSVVFTRETGMTPSQWRHLNSQK
                     D"
     CDS             complement(4989..5825)
                     /codon_start=1
                     /gene="E. coli rhaS"
                     /product="positive regulator of the rhaB promoter
                     "
                     /label=rhaS
                     /note="activated by L-rhamnose"
                     /translation="MTVLHSVDFFPSGNASVAIEPRLPQADFPEHHHDFHEIVIVEHGT
                     GIHVFNGQPYTITGGTVCFVRDHDRHLYEHTDNLCLTNVLYRSPDRFQFLAGLNQLLPQ
                     ELDGQYPSHWRVNHSVLQQVRQLVAQMEQQEGENDLPSTASREILFMQLLLLLRKSSLQ
                     ENLENSASRLNLLLAWLEDHFADEVNWDAVADQFSLSLRTLHRQLKQQTGLTPQRYLNR
                     LRLMKARHLLRHSEASVTDIAYRCGFSDSNHFSTLFRREFNWSPRDIRQGRDGFLQ"
     promoter        5971..6089
                     /label=rhaB promoter
                     /note="promoter of the E. coli rhaBAD operon, conferring 
                     tight induction with L-rhamnose and repression with 
                     D-glucose in the presence of RhaR and RhaS (Giacalone et 
                     al., 2006)"
ORIGIN
        1 ggcggctaca gccgatagtc tggaacagcg cacttacggg ttgctgcgca acccaagtgc
       61 taccggcgcg gcagcgtgac ccgtgtcggc ggctccaacg gctcgccatc gtccagaaaa
      121 cacggctcat cgggcatcgg caggcgctgc tgcccgcgcc gttcccattc ctccgtttcg
      181 gtcaaggctg gcaggtctgg ttccatgccc ggaatgccgg gctggctggg cggctcctcg
      241 ccggggccgg tcggtagttg ctgctcgccc ggatacaggg tcgggatgcg gcgcaggtcg
      301 ccatgcccca acagcgattc gtcctggtcg tcgtgatcaa ccaccacggc ggcactgaac
      361 accgacaggc gcaactggtc gcggggctgg ccccacgcca cgcggtcatt gaccacgtag
      421 gccgacacgg tgccggggcc gttgagcttc acgacggaga tccagcgctc ggccaccaag
      481 tccttgactg cgtattggac cgtccgcaaa gaacgtccga tgagcttgga aagtgtcttc
      541 tggctgacca ccacggcgtt ctggtggccc atctgcgcca cgaggtgatg cagcagcatt
      601 gccgccgtgg gtttcctcgc aataagcccg gcccacgcct catgcgcttt gcgttccgtt
      661 tgcacccagt gaccgggctt gttcttggct tgaatgccga tttctctgga ctgcgtggcc
      721 atgcttatct ccatgcggta ggggtgccgc acggttgcgg caccatgcgc aatcagctgc
      781 aacttttcgg cagcgcgaca acaattatgc gttgcgtaaa agtggcagtc aattacagat
      841 tttctttaac ctacgcaatg agctattgcg gggggtgccg caatgagctg ttgcgtaccc
      901 ccctttttta agttgttgat ttttaagtct ttcgcatttc gccctatatc tagttctttg
      961 gtgcccaaag aagggcaccc ctgcggggtt cccccacgcc ttcggcgcgg ctccccctcc
     1021 ggcaaaaagt ggcccctccg gggcttgttg atcgactgcg cggccttcgg ccttgcccaa
     1081 ggtggcgctg cccccttgga acccccgcac tcgccgccgt gaggctcggg gggcaggcgg
     1141 gcgggcttcg cccttcgact gcccccactc gcataggctt gggtcgttcc aggcgcgtca
     1201 aggccaagcc gctgcgcggt cgctgcgcga gccttgaccc gccttccact tggtgtccaa
     1261 ccggcaagcg aagcgcgcag gccgcaggcc ggaggctttt ccccagagaa aattaaaaaa
     1321 attgatgggg caaggccgca ggccgcgcag ttggagccgg tgggtatgtg gtcgaaggct
     1381 gggtagccgg tgggcaatcc ctgtggtcaa gctcgtgggc aggcgcagcc tgtccatcag
     1441 cttgtccagc agggttgtcc acgggccgag cgaagcgagc cagccggtgg ccgctcgcgg
     1501 ccatcgtcca catatccacg ggctggcaag ggagcgcagc gaccgcgcag ggcgaagccc
     1561 ggagagcaag cccgtagggc gccgcagccg ccgtaggcgg tcacgacttt gcgaagcaaa
     1621 gtctagtgag tatactcaag cattgagtgg cccgccggag gcaccgcctt gcgctgcccc
     1681 cgtcgagccg gttggacacc aaaagggagg ggcaggcatg gcggcatacg cgatcatgcg
     1741 atgcaagaag ctggcgaaaa tgggcaacgt ggcggccagt ctcaagcacg cctaccgcga
     1801 gcgcgagacg cccaacgctg acgccagcag gacgccagag aacgagcact gggcggccag
     1861 cagcaccgat gaagcgatgg gccgactgcg cgagttgctg ccagagaagc ggcgcaagga
     1921 cgctgtgttg gcggtcgagt acgtcatgac ggccagcccg gaatggtgga agtcggccag
     1981 ccaagaacag caggcggcgt tcttcgagaa ggcgcacaag tggctggcgg acaagtacgg
     2041 ggcggatcgc atcgtgacgg ccagcatcca ccgtgacgaa accagcccgc acatgaccgc
     2101 gttcgtggtg ccgctgacgc aggacggcag gctgtcggcc aaggagttca tcggcaacaa
     2161 agcgcagatg acccgcgacc agaccacgtt tgcggccgct gtggccgatc tagggctgca
     2221 acggggcatc gagggcagca aggcacgtca cacgcgcatt caggcgttct acgaggccct
     2281 ggagcggcca ccagtgggcc acgtcaccat cagcccgcaa gcggtcgagc cacgcgccta
     2341 tgcaccgcag ggattggccg aaaagctggg aatctcaaag cgcgttgaga cgccggaagc
     2401 cgtggccgac cggctgacaa aagcggttcg gcaggggtat gagcctgccc tacaggccgc
     2461 cgcaggagcg cgtgagatgc gcaagaaggc cgatcaagcc caagagacgg cccgagacct
     2521 tcgggagcgc ctgaagcccg ttctggacgc cctggggccg ttgaatcggg atatgcaggc
     2581 caaggccgcc gcgatcatca aggccgtggg cgaaaagctg ctgacggaac agcgggaagt
     2641 ccagcgccag aaacaggccc agcgccagca ggaacgcggg cgcgcacatt tccccgaaaa
     2701 gtgccacctg ggatgaatgt cagctactgg gctatctgga caagggaaaa cgcaagcgca
     2761 aagagaaagc aggtagcttg cagtgggctt acatggcgat agctagactg ggcggtttta
     2821 tggacagcaa gcgaaccgga attgccagct ggggcgccct ctggtaaggt tgggaagccc
     2881 tgcaaagtaa actggatggc tttcttgccg ccaaggatct gatggcgcag gggatcaaga
     2941 tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     3001 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     3061 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     3121 aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg
     3181 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     3241 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     3301 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     3361 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     3421 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     3481 ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc
     3541 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     3601 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     3661 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     3721 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     3781 ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg
     3841 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     3901 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc ccatgggcaa atattatacg
     3961 caaggcgaca aggtgctgat gccgctggcg attcaggttc atcatgccgt ttgtgatggc
     4021 ttccatgtcg gcagaatgct taatgaatta caacagtttt tatgcattaa tctttctgcg
     4081 aattgagatg acgccactgg ctgggcgtca tcccggtttc ccgggtaaac accaccgaaa
     4141 aatagttact atcttcaaag ccacattcgg tcgaaatatc actgattaac aggcggctat
     4201 gctggagaag atattgcgca tgacacactc tgacctgtcg cagatattga ttgatggtca
     4261 ttccagtctg ctggcgaaat tgctgacgca aaacgcgctc actgcacgat gcctcatcac
     4321 aaaatttatc cagcgcaaag ggacttttca ggctagccgc cagccgggta atcagcttat
     4381 ccagcaacgt ttcgctggat gttggcggca acgaatcact ggtgtaacga tggcgattca
     4441 gcaacatcac caactgcccg aacagcaact cagccatttc gttagcaaac ggcacatgct
     4501 gactactttc atgctcaagc tgaccgataa cctgccgcgc ctgcgccatc cccatgctac
     4561 ctaagcgcca gtgtggttgc cctgcgctgg cgttaaatcc cggaatcgcc ccctgccagt
     4621 caagattcag cttcagacgc tccgggcaat aaataatatt ctgcaaaacc agatcgttaa
     4681 cggaagcgta ggagtgttta tcgtcagcat gaatgtaaaa gagatcgcca cgggtaatgc
     4741 gataagggcg atcgttgagt acatgcaggc cattaccgcg ccagacaatc accagctcac
     4801 aaaaatcatg tgtatgttca gcaaagacat cttgcggata acggtcagcc acagcgactg
     4861 cctgctggtc gctggcaaaa aaatcatctt tgagaagttt taactgatgc gccaccgtgg
     4921 ctacctcggc cagagaacga agttgattat tcgcaatatg gcgtacaaat acgttgagaa
     4981 gattcgcgtt attgcagaaa gccatcccgt ccctggcgaa tatcacgcgg tgaccagtta
     5041 aactctcggc gaaaaagcgt cgaaaagtgg ttactgtcgc tgaatccaca gcgataggcg
     5101 atgtcagtaa cgctggcctc gctgtggcgt agcagatgtc gggctttcat cagtcgcagg
     5161 cggttcaggt atcgctgagg cgtcagtccc gtttgctgct taagctgccg atgtagcgta
     5221 cgcagtgaaa gagaaaattg atccgccacg gcatcccaat tcacctcatc ggcaaaatgg
     5281 tcctccagcc aggccagaag caagttgaga cgtgatgcgc tgttttccag gttctcctgc
     5341 aaactgcttt tacgcagcaa gagcagtaat tgcataaaca agatctcgcg actggcggtc
     5401 gagggtaaat cattttcccc ttcctgctgt tccatctgtg caaccagctg tcgcacctgc
     5461 tgcaatacgc tgtggttaac gcgccagtga gacggatact gcccatccag ctcttgtggc
     5521 agcaactgat tcagcccggc gagaaactga aatcgatccg gcgagcgata cagcacattg
     5581 gtcagacaca gattatcggt atgttcatac agatgccgat catgatcgcg tacgaaacag
     5641 accgtgccac cggtgatggt atagggctgc ccattaaaca catgaatacc cgtgccatgt
     5701 tcgacaatca caatttcatg aaaatcatga tgatgttcag gaaaatccgc ctgcgggagc
     5761 cggggttcta tcgccacgga cgcgttacca gacggaaaaa aatccacact atgtaatacg
     5821 gtcatactgg cctcctgatg tcgtcaacac ggcgaaatag taatcacgag gtcaggttct
     5881 taccttaaat tttcgacgga aaaccacgta aaaaacgtcg atttttcaag atacagcgtg
     5941 aattttcagg aaatgcggtg agcatcacat caccacaatt cagcaaattg tgaacatcat
     6001 cacgttcatc tttccctggt tgccaatggc ccattttcct gtcagtaacg agaaggtcgc
     6061 gaattcaggc gctttttaga ctggtcgtaa tgaacaattc ttaagaagga gatatacata
     6121 tgcgaatcgt tgaatgcgca cagggtagcg acgaatggca tcgcgcgcgt gccggcgtca
     6181 tcacggccag catgttcgcc gcggcgcgca agcgtctcaa ggtcggccct aacaagggcg
     6241 attacacgtc cgaggcgaaa gactatgcat tccgcctcgc ggtcgagcgc attggcggtc
     6301 agccgctcga cgaaggattc gagacgtggc agatgaagcg cgggcacgag ctcgaacccg
     6361 aggcgcgcat ggagcacgaa gcactcacag gcctgttcgt gaagcgcgcc ggattcgtcg
     6421 tgaccgatga cggactgttc ggcgcgagcg ctgacggcct gatcgaaccc gatggcggcg
     6481 ccgaatacaa atgcttcgtt gcgccggaca agctgcgccc gatgctgctc gacaacgacg
     6541 ccggcgacgt catggaacag gcgcagggct gcatgtggct ttgcggcgcg aagtggtggc
     6601 acgtcggcct gtactgcccc gccctcgccg ctgctggccg ccagttcacg ctgatcaagg
     6661 tacaacgcga cgatgcattc atcgagcgca tggaggccga tctcgtcgag ttctcgcgtc
     6721 tcgtcactgc gtacgaaaca cggctgcgcc agcacccagc cattgaggct tgaaggaggc
     6781 agctatatga gcaatgccct catcgccgtc accaacgaaa tctacggcgt ccgtgaaaac
     6841 ttcgagcgtg tcgccgtcga tcgctcgatc aacttcgacc gcgaggccgg atttgcagtg
     6901 cagatcctgc aaggcaatcc gtacttgctc ggcgtcgccg agaaggatcg cgcttcgatg
     6961 atcgcggcgg tgacgaacgt ttcggcgatt ggcatcagcc tcaaccccgc gaagaagcag
     7021 gcgtacctcg tgccgcgtgg cggcaaggtc tgtctcgaca tcagctacat ggggctgctg
     7081 gatctggcga tcgcctccgg ttcgatccag tggggtcaag ccgaagtggt gtgcgaagaa
     7141 gacacgttca ccctgcgcgg cttcgacaaa gcgccgttgc atgacttcgc accgttcagc
     7201 aagacgcgtg gcgcagtggt cggcgtctat gtcgtcgtca agaccgcgac cggcgactac
     7261 ctgaccgaca cgatgacgat cgccgaggta caggcgatca aggctcgctc cgaaaccggc
     7321 aagaaggaca ccgggccgtg gaaaacggac ttcaacgaaa tggcgaagaa gaccgtcatc
     7381 aagcgagcct acaagacgtg gccgaagacg gatcgccttg atgcggccgt gcaccacctg
     7441 aacaccgatg gcggcgaagg catcaacttc gccagtgagc gcccgaatta ctgcgacccg
     7501 gacgtgctga aagcatgggt cgacaaggcg aaggctgcgc gcaccgctga atcgctcgcg
     7561 aagatttggc aggacggttt ggctgtcatc cggcctacca aagacatgca gacttacgaa
     7621 gtattcaagc agacggtcgc ggttcgtggc gaagaaatta agcgccgcgc cgagcagcgg
     7681 aacaatgctc aggacgtgac gccgactgag ccagcacagc gtgagccggg cgccgacgac
     7741 gagctcgaag aagatttcag gcgccagatg gagcgcgagc aagggggtgc cgatgcgaat
     7801 cgttgactcg aggaggggat cctatgaacc cgatcaccta cctgtgcggc gcgctcgacc
     7861 ggctgttcga acgtagtccg gtcgcgggga ttgttgttgc gatcgtgatt gcggccggat
     7921 gcgtgatcgc gatcgcctat ctcaaccagg acggcgagtc cgtcaccact atcaatgcga
     7981 ggtttgcatg actcgaggat cctgtacacc cagcccgcct aatgagcggg cttttttttg
     8041 aacaaagctt accggtttat tgactaccgg aagcagtgtg accgtgtgct tctcaaatgc
     8101 ctgaggccag tttgctcagg ctctccccgt ggaggtaata attgacgata tgatcattta
     8161 ttctgcctcc cagagcctga taaaaacggt gaatccgtta gcgaggtgcc gccggcttcc
     8221 attcaggtcg aggtggcccg gctccatgca ccgcgacgca acgcggggag gcagacaagg
     8281 tatagggcgg cga
//
