LOCUS       sequence-225649-        7028 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway compatible allelic exchange vector for Burkholderia sp..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7028)
  AUTHORS   Fazli M, Harrison JJ, Gambino M, Givskov M, Tolker-Nielsen T
  TITLE     In-Frame and Unmarked Gene Deletions in Burkholderia cenocepacia via
            an Allelic Exchange System Compatible with Gateway Technology.
  JOURNAL   Appl Environ Microbiol. 2015 Jun;81(11):3623-30. doi: 
            10.1128/AEM.03909-14. Epub 2015 Mar 20.
  PUBMED    25795676
REFERENCE   2  (bases 1 to 7028)
  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: 
            "10.1128/AEM.03909-14"; journalName: "Appl Environ Microbiol"; date:
            "2015-06"; volume: "81"; issue: "11"; pages: "3623-30"
COMMENT     Sequence Label: pDONRPEX18Tp-SceI-pheS
FEATURES             Location/Qualifiers
     source          1..7028
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     40..58
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(96..118)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     218..237
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     oriT            1508..1617
                     /note="incP origin of transfer"
     terminator      complement(2060..2087)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(2179..2265)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     2418..2435
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     primer_bind     2615..2631
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2688..2704
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2721..2952
                     /gene="mutant version of attP"
                     /label=attP1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(3348..3653)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(3450..3469)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(3973..4656)
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGRKRR
                     VDPAY"
     primer_bind     4571..4590
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4657..4759)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     protein_bind    complement(4904..5135)
                     /gene="mutant version of attP"
                     /label=attP2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(5153..5172)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5154..5172)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5177..5193)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5266..5282)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5279..5301)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5290..5306
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5314..5320,5321..5338,5339..5344))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5359..5380
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5668..6256)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5748..5767)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        join(6374..6379,6380..6396,6397..6402)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             6728..6964
                     /codon_start=1
                     /gene="dhfrIIb from plasmid R388"
                     /product="E. coli plasmid-associated dihydrofolate 
                     reductase"
                     /label=TpR
                     /note="confers resistance to trimethoprim"
                     /translation="MGQSSDEANAPVAGQFALPLSATFGLGDRVRKKSGAAWQGQVVGW
                     YCTKLTPEGYAVESESHPGSVQIYPVAALERVA"
ORIGIN
        1 gtttgacagc ttatcatcga catgacatta acctataaaa ataggcgtat cacgaggccc
       61 tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag
      121 acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca
      181 gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg catcagagca gattgtactg
      241 agagtgcacc ataatcggca ttttcttttg cgtttttatt tgttaactgt taattgtcct
      301 tgttcaagga tgctgtcttt gacaacagat gttttcttgc ctttgatgtt cagcaggaag
      361 ctaggcgcaa acgttggatc tttaggcgaa ctggcggagg aaccggaggt cattttcgaa
      421 aaagagccgg aggtcctgga caccataccg gagcatggta aggcgttcca acccggaacc
      481 gaagccgaag ccaatgtaac gctccgggtc gaggcccata ttccgaatga ccgtggggtg
      541 gacctgaccg ctgccgctga tttccaacca cttaccagca ttcttcccat gttcaaacat
      601 catatcaatt tccgcgctgg gttccgtgaa ggggaagtag ctcgggcgga aacggaccaa
      661 gatgtcatct cgttcgaaaa atttcttgag aaagtcggtg taggcaccct tgagatcggc
      721 aaacgacacg ttttcatcaa tccagagccc ctccacctga ttaaacatgg ggctatgggt
      781 ggcgtccgaa tcgacccgat aggtccgacc gggagcaatc accttaatcg ggggtcgatt
      841 catgcgggcg taccgcacct gcatgggcga ggtatgggtc cggaggagga ggggccgccc
      901 atcggcgtcc ttaccatcca catagaaggt gtcctgcatg gaccgtgcgg ggtggttttc
      961 gggcgagttg agcgccgtga aattatacca atccgtctca atttccggtc catccgcgac
     1021 atcgaagcca atgctacgga aaatctgctc cacacgttcc caggttcgca tgacggggtg
     1081 cagcgagccg gcgcccgccc cgcgaccggg gagggtcaca tcaatggctt cagccgccag
     1141 gcgctgattg agaagggcat ccgccaaggc ctgccgccgg gcattcaagg ccgcctccac
     1201 ctgctgcttg gccacgttaa tccgggcacc ctcggtcttc cgggtttccg ggtccagctt
     1261 acccagcccc ttcaagagtt ccgtgagggc accgctcttg cccaggaagc gggccttttc
     1321 attttccaag gtcgtaatgt cggccgcgcc ctcaaagctc tgctgggcat ccgccacgat
     1381 ctgatcgagg tccatatgct tgtcctttcg aagctcgatc cgattgtcag acccggcatg
     1441 atattccgaa aatcccaagc gagtcaacag ctagtgcttg ccctcagtac tgttacgccg
     1501 gcggtagccg gccagcctcg cagagcagga ttcccgttga gcaccgccag gtgcgaataa
     1561 gggacagtga agaaggaaca cccgctcgcg ggtgggccta cttcacctat cctgcccggc
     1621 tgacgccgtt ggatacacca aggaaagtct acacgaaccc tttggcaaaa tcctgtatat
     1681 cgtgcgaaaa aggatggata taccgaaaaa atcgctataa tgaccccgaa gcagggttat
     1741 gcagcggaaa agcgctgctt ccctgctgtt ttgtggaata tctaccgact ggaaacaggc
     1801 aaatgcagga aattactgaa ctgaggggac aggcgagaga cgatgccaaa gagctacacc
     1861 gacgagctgg ccgagtgggt tgaatcccgc gcggccaaga agcgccggcg tgatgaggct
     1921 gcggttgcgt tcctggcggt gagggcggat gtcgatatgc ggtgtgaaat accgcacaga
     1981 tgcgtaagga gaaaataccg catcaggcgc atatttgaat gtatttagaa aaataaacaa
     2041 aagagtttgt agaaacgcaa aaaggccatc cgtcaggatg gccttctgct taatttgatg
     2101 cctggcagtt tatggcgggc gtcctgcccg ccaccctccg ggccgttgct tcgcaacgtt
     2161 caaatccgct cccggcggat ttgtcctact caggagagcg ttcaccgaca aacaacagat
     2221 aaaacgaaag gcccagtctt tcgactgagc ctttcgtttt atttgatgcc tggcagttcc
     2281 ctactctcgc atggggagac cccacactac catcggcgct acggcgtttc acttctgagt
     2341 tcggcatggg gtcaggtggg accaccgcgc tactgccgcc aggcaaattc tgttttatca
     2401 gaccgcttct gcgttctgat ttaatctgta tcaggctgaa aatcttctct catccgccaa
     2461 aacagccaag ctcgccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg
     2521 cgggcctctt cgctattacg ccagctggcg aaagggggat gtgctgcaag gcgattaagt
     2581 tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa cgacggccag tgccaagctt
     2641 tagggataac agggtaatag catggatgtt ttcccagtca cgacgttgta aaacgacggc
     2701 cagtcttaag ctcgggcccc aaataatgat tttattttga ctgatagtga cctgttcgtt
     2761 gcaacaaatt gatgagcaat gcttttttat aatgccaact ttgtacaaaa aagctgaacg
     2821 agaaacgtaa aatgatataa atatcaatat attaaattag attttgcata aaaaacagac
     2881 tacataatac tgtaaaacac aacatatcca gtcactatga atcaactact tagatggtat
     2941 tagtgacctg tagtcgaccg acagccttcc aaatgttctt cgggtgatgc tgccaactta
     3001 gtcgaccgac agccttccaa atgttcttct caaacggaat cgtcgtatcc agcctactcg
     3061 ctattgtcct caatgccgta ttaaatcata aaaagaaata agaaaaagag gtgcgagcct
     3121 cttttttgtg tgacaaaata aaaacatcta cctattcata tacgctagtg tcatagtcct
     3181 gaaaatcatc tgcatcaaga acaatttcac aactcttata cttttctctt acaagtcgtt
     3241 cggcttcatc tggattttca gcctctatac ttactaaacg tgataaagtt tctgtaattt
     3301 ctactgtatc gacctgcaga ctggctgtgt ataagggagc ctgacattta tattccccag
     3361 aacatcaggt taatggcgtt tttgatgtca ttttcgcggt ggctgagatc agccacttct
     3421 tccccgataa cggagaccgg cacactggcc atatcggtgg tcatcatgcg ccagctttca
     3481 tccccgatat gcaccaccgg gtaaagttca cgggagactt tatctgacag cagacgtgca
     3541 ctggccaggg ggatcaccat ccgtcgcccg ggcgtgtcaa taatatcact ctgtacatcc
     3601 acaaacagac gataacggct ctctctttta taggtgtaaa ccttaaactg catttcacca
     3661 gcccctgttc tcgtcagcaa aagagccgtt catttcaata aaccgggcga cctcagccat
     3721 cccttcctga ttttccgctt tccagcgttc ggcacgcaga cgacgggctt cattctgcat
     3781 ggttgtgctt accagaccgg agatattgac atcatatatg ccttgagcaa ctgatagctg
     3841 tcgctgtcaa ctgtcactgt aatacgctgc ttcatagcat acctcttttt gacatacttc
     3901 gggtatacat atcagtatat attcttatac cgcaaaaatc agcgcgcaaa tacgcatact
     3961 gttatctggc ttttagtaag ccggatccac gcggcgttta cgccccccct gccactcatc
     4021 gcagtactgt tgtaattcat taagcattct gccgacatgg aagccatcac aaacggcatg
     4081 atgaacctga atcgccagcg gcatcagcac cttgtcgcct tgcgtataat atttgcccat
     4141 ggtgaaaacg ggggcgaaga agttgtccat attggccacg tttaaatcaa aactggtgaa
     4201 actcacccag ggattggctg agacgaaaaa catattctca ataaaccctt tagggaaata
     4261 ggccaggttt tcaccgtaac acgccacatc ttgcgaatat atgtgtagaa actgccggaa
     4321 atcgtcgtgg tattcactcc agagcgatga aaacgtttca gtttgctcat ggaaaacggt
     4381 gtaacaaggg tgaacactat cccatatcac cagctcaccg tctttcattg ccatacggaa
     4441 ttccggatga gcattcatca ggcgggcaag aatgtgaata aaggccggat aaaacttgtg
     4501 cttatttttc tttacggtct ttaaaaaggc cgtaatatcc agctgaacgg tctggttata
     4561 ggtacattga gcaactgact gaaatgcctc aaaatgttct ttacgatgcc attgggatat
     4621 atcaacggtg gtatatccag tgattttttt ctccatttta gcttccttag ctcctgaaaa
     4681 tctcgataac tcaaaaaata cgcccggtag tgatcttatt tcattatggt gaaagttgga
     4741 acctcttacg tgccgatcaa cgtctcattt tcgccaaaag ttggcccagg gcttcccggt
     4801 atcaacaggg acaccaggat ttatttattc tgcgaagtga tcttccgtca caggtattta
     4861 ttcggcgcaa agtgcgtcgg gtgatgctgc caacttagtc gactacaggt cactaatacc
     4921 atctaagtag ttgattcata gtgactggat atgttgtgtt ttacagtatt atgtagtctg
     4981 ttttttatgc aaaatctaat ttaatatatt gatatttata tcattttacg tttctcgttc
     5041 agctttcttg tacaaagttg gcattataag aaagcattgc ttatcaattt gttgcaacga
     5101 acaggtcact atcagtcaaa ataaaatcat tatttgccat ccagctgata tcccctatag
     5161 tgagtcgtat tacatggtca tagctgtttc ctggcagctc tggcccgtgt ctcaaaatct
     5221 ctgatctaga ggatccccgg gtaccgagct cgaattcgta atcatgtcat agctgtttcc
     5281 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     5341 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     5401 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5461 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgacc cgctgcgctc
     5521 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5581 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5641 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5701 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5761 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5821 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5881 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5941 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     6001 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     6061 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     6121 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     6181 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     6241 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     6301 cgaaaactca cgttaaggga ttttggtcat gacgcactag tggaacgaaa tcgatgagct
     6361 cgcacgaacc cagttgacat aagcctgttc ggttcgtaaa ctgtaatgca agtagcgtat
     6421 gcgctcacgc aactggtcca gaaccttgac cgaacgcagc ggtggtaacg gcgcagtggc
     6481 ggttttcatg gcttgttatg actgtttttt tgtacagtct atgcctcggg catccaagta
     6541 gcaagcgcgt tacgccgtgg gtcgatgttt gatgttatgg agcagcaacg atgttacgca
     6601 gcagggcagt cgccctaaaa caaagttagg cagccgttgt gctggtgctt tctgatagtt
     6661 gttgtggggt aggcagtcag agctcgattt gcttgtcgcc ataatagatt cacaagaagg
     6721 attcgacatg ggtcaaagta gcgatgaagc caacgctccc gttgcagggc agtttgcgct
     6781 tcccctgagt gccacctttg gcttagggga tcgcgtacgc aagaaatctg gtgccgcttg
     6841 gcagggtcaa gtcgtcggtt ggtattgcac aaaactcact cctgaaggct atgcggtcga
     6901 gtccgaatcc cacccaggct cagtgcaaat ttatcctgtg gctgcacttg aacgtgtggc
     6961 ctaagcgata tcttaggaks ysccrkygrt gmyggccacg atgcgtccgg cgtagaggat
     7021 ctgctcat
//
