LOCUS       Exported                7658 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Derived from mini-CTX1. An integrase vector that can integrate the 
            rhamnose-inducible promoter system into the chromosome of a host 
            bacterium..
ACCESSION   .
VERSION     .
KEYWORDS    pAH-CTX1-rha
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7658)
  AUTHORS   Hogan AM, Rahman ASMZ, Lightly TJ, Cardona ST
  TITLE     A broad-host-range CRISPRi toolkit for silencing gene expression in 
            Burkholderia.
  JOURNAL   ACS Synth Biol. 2019 Sep 6. doi: 10.1021/acssynbio.9b00232.
  PUBMED    31491085
REFERENCE   2  (bases 1 to 7658)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7658
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    complement(153..200)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     737..757
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        739..757
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        complement(822..940)
                     /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)"
     CDS             1086..1922
                     /codon_start=1
                     /gene="E. coli rhaS"
                     /product="positive regulator of the rhaB promoter
                     "
                     /label=rhaS
                     /note="activated by L-rhamnose"
                     /translation="MTVLHSVDFFPSGNASVAIEPRLPQADFPEHHHDFHEIVIVEHGT
                     GIHVFNGQPYTITGGTVCFVRDHDRHLYEHTDNLCLTNVLYRSPDRFQFLAGLDQLLPQ
                     ELDGQYPSHWRVNHSVLQQVRQLVAQMEQQEGENDLPSTASREILFMQLLLLLRKSSLQ
                     ENLENSASRLNLLLAWLEDHFADEVNWDAVADQFSLSLRTLHRQLKQQTGLTPQRYLNR
                     LRLMKARHLLRHSEASVTDIAYRCGFSDSNHFSTLFRREFNWSPRDIRQGRDGFLQ"
     CDS             1996..2844
                     /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"
     primer_bind     complement(2888..2904)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2889..2905)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2929..2948)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2930..2948)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    complement(3122..3169)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     oriT            3376..3484
                     /note="incP origin of transfer"
     rep_origin      complement(4916..5504)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4996..5015)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6326..7516)
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     7145..7164
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        complement(7564..7592)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
ORIGIN
        1 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
       61 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
      121 ttaatgcagc gaattagctt caaaagcgct ctgaagttcc tatactttct agagaatagg
      181 aacttcggaa taggaacttc aagatcccct gattcccttt gtcaacagca atggatttca
      241 atgggtcaac atgaaatccg gctgttgccc tgggacggca ggtttgacgg caggcttcca
      301 ggctctctcg cacacgctgg gaacactctg ggaacaaacg cgagtgaaaa tggacagtag
      361 ggcccaggaa taatgcggcc tccagcgagg ttggggcgtt tcatatgagt tcgaatctca
      421 ccgcctccgc catataacct gctgttttca ttgatgtttc cattcgaccc actctcagga
      481 gtgaacgatt tgggaacaaa ctgggaacga ggcgcagaaa aaaggggccc cgcagggccc
      541 ctttttcgtt gcttcaggaa agtttcagcg cctgattcag cagacctacg atgtcggggc
      601 cgtccttgct gaattagctt tatgcttgta aaccgttttg tgaaaaaatt tttaaaataa
      661 aaaaggggac ctctagggtc cccaattaat tagtaatata atctattaaa ggtcattcaa
      721 aaggtcatcc acccgcgcaa ttaaccctca ctaaagggaa caaaagctgg agctccaccg
      781 cggtggcggc cgctctagaa ctagttcctg ctgaatttca ttacgaccag tctaaaaagc
      841 gcctgaattc gcgaccttct cgttactgac aggaaaatgg gccattggca accagggaaa
      901 gatgaacgtg atgatgttca caatttgctg aattgtggtg atgtgatgct caccgcattt
      961 cctgaaaatt cacgctgtat cttgaaaaat cgacgttttt tacgtggttt tccgtcgaaa
     1021 atttaaggta agaacctgac ctcgtgatta ctatttcgcc gtgttgacga catcaggagg
     1081 ccagtatgac cgtattacat agtgtggatt tttttccgtc tggtaacgcg tccgtggcga
     1141 tagaaccccg gctcccgcag gcggattttc ctgaacatca tcatgatttt catgaaattg
     1201 tgattgtcga acatggcacg ggtattcatg tgtttaatgg gcagccctat accatcaccg
     1261 gtggcacggt ctgtttcgta cgcgatcatg atcggcatct gtatgaacat accgataatc
     1321 tgtgtctgac caatgtgctg tatcgctcgc cggatcgatt tcagtttctc gccgggctgg
     1381 atcagttgct gccacaagag ctggatgggc agtatccgtc tcactggcgc gttaaccaca
     1441 gcgtattgca gcaggtgcga cagctggttg cacagatgga acagcaggaa ggggaaaatg
     1501 atttaccctc gaccgccagt cgcgagatct tgtttatgca attactgctc ttgctgcgta
     1561 aaagcagttt gcaggagaac ctggaaaaca gcgcatcacg tctcaacttg cttctggcct
     1621 ggctggagga ccattttgcc gatgaggtga attgggatgc cgtggcggat caattttctc
     1681 tttcactgcg tacgctacat cggcagctta agcagcaaac gggactgacg cctcagcgat
     1741 acctgaaccg cctgcgactg atgaaagccc gacatctgct acgccacagc gaggccagcg
     1801 ttactgacat cgcctatcgc tgtggattca gcgacagtaa ccacttttcg acgctttttc
     1861 gccgagagtt taactggtca ccgcgtgata ttcgccaggg acgggatggc tttctgcaat
     1921 aacgcgaatc ttctcaacgt atttgtacgc catattgcga ataatcaact tcgttctctg
     1981 gccgaggtag ccacggtggc gcatcagtta aaacttctca aagatgattt ttttgccagc
     2041 gaccagcagg cagtcgctgt ggctgaccgt tatccgcaag atgtctttgc tgaacataca
     2101 catgattttt gtgagctggt gattgtctgg cgcggtaatg gcctgcatgt actcaacgat
     2161 cgcccttatc gcattacccg tggcgatctc ttttacattc atgctgacga taaacactcc
     2221 tacgcttccg ttaacgatct ggttttgcag aatattattt attgcccgga gcgtctgaag
     2281 ctgaatcttg actggcaggg ggcgattccg ggatttaacg ccagcgcagg gcaaccacac
     2341 tggcgcttag gtagcatggg gatggcgcag gcgcggcagg ttatcggtca gcttgagcat
     2401 gaaagtagtc agcatgtgcc gtttgctaac gaaatggctg agttgctgtt cgggcagttg
     2461 gtgatgttgc tgaatcgcca tcgttacacc agtgattcgt tgccgccaac atccagcgaa
     2521 acgttgctgg ataagctgat tacccggctg gcggctagcc tgaaaagtcc ctttgcgctg
     2581 gataaatttt gtgatgaggc atcgtgcagt gagcgcgttt tgcgtcagca atttcgccag
     2641 cagactggaa tgaccatcaa tcaatatctg cgacaggtca gagtgtgtca tgcgcaatat
     2701 cttctccagc atagccgcct gttaatcagt gatatttcga ccgaatgtgg ctttgaagat
     2761 agtaactatt tttcggtggt gtttacccgg gaaaccggga tgacgcccag ccagtggcgt
     2821 catctcaatt cgcagaaaga ttaaatgcat cgatgagctc gcacgaaccc agttgacata
     2881 agcttatcga taccgtcgac ctcgaggggg ggcccggtac ccaattcgcc ctatagtgag
     2941 tcgtattacg cgcgccggtg gatgaccttt tgaatgacct ttaatagatt atattactaa
     3001 ttaattgggg accctagagg tccccttttt tattttaaaa attttttcac aaaacggttt
     3061 acaagcataa agctaattcc attgagtaag tttttaagca catcagcttc aaaagcgctc
     3121 tgaagttcct atactttcta gagaatagga acttcggaat aggtacttca agatccccaa
     3181 ttcgctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     3241 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     3301 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     3361 catgctagag cttatcggcc agcctcgcag agcaggattc ccgttgagca ccgccaggtg
     3421 cgaataaggg acagtgaaga aggaacaccc gctcgcgggt gggcctactt cacctatcct
     3481 gcccggctga cgccgttgga tacaccaagg aaagtctaca cgaacccttt ggcaaaatcc
     3541 tgtatatcgt gcgaaaaagg atggatatac cgaaaaaatc gctataatga ccccgaagca
     3601 gggttatgca gcggaaagta taccttaacc tttttcgttg cttcaggaaa gtttcagcgc
     3661 ctgattcagc agacctacga tgtcggggcc gtccttgctg atccacttgg cgtaatgctt
     3721 gaagatcatc gccgtcgagg tgtgccccat ctggtccgcg atccattcgg gagtggcgat
     3781 gccgctgctg agcatctggc tggcgaaggt gtggcggcac tgattgggac cacgcgagcg
     3841 gacgcccgca ttcctgaggt ggccatgcca ccagccgttg cgcagcgtat ccgacgtgga
     3901 gtacgccgcc ccgctagcac tgttgtggaa cacgaagcgt acccgttgag gctttctggt
     3961 ccggttgtcg cggtctatca cctcgatctg tacgggcgga aggcgtctgg ttagcttggc
     4021 ctgctgctgg agtgcgcgca gcgccggggc cagtagctgg accttcctcg tcgatcggcg
     4081 agtcttggtg accttgtact ggctgcgtac acgagcgcgg cggaaaacca cggtccccgt
     4141 gtccaaatcg acatcttccc aggccagagc tatggcctcg ctgacgcggg ggccgctcca
     4201 gatcatgaac tccgccaggt tcagctcacc gatgcgcgcg gtttcagtac cgaggatgag
     4261 gtcgatttcc tcacgagtga atgggtctgg gtcatccgca tccggaagag tgatcacgat
     4321 gccgtctgtt gggtcgtgcg cgctccggtt ccgagttcga tacagccgga agatctgccg
     4381 caggttgctg acgatctctc gcaccgtctt gttgtgcagc ttgggcatga gcgtgttctg
     4441 cacccagtct tgaatatcca ggtggtcgat gctgtcggcc tggctgtcac cccagcgagg
     4501 tcggatgtgg gtttccacgc ggctggtgta gccgcggaag ccgctggcgg caatctggtt
     4561 gcgtttgatg tcgagccaga ggtcgatgta atggcccaag gtgttgcttt tcacccgcgg
     4621 tgagtccggg aagtgtcggc tgtagctgaa gactccttgc ttgatctcgt agttgatgat
     4681 gccggccagc tgttcggcat tggctatatt ctcgggcgtt gcgtcccctg gtatggactc
     4741 cctacacagc tcgccctggt agcggaagta gatcctgatc cgttttccgc gtacctcgac
     4801 tccgtctgcc atctggttcc cctgatccac accaagcgtt caggctacca agctggatgt
     4861 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4921 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     4981 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     5041 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     5101 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5161 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5221 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5281 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5341 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5401 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5461 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5521 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5581 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5641 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     5701 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5761 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     5821 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5881 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     5941 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     6001 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6061 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6121 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6181 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtca agggttggtt
     6241 tgcgcattca cagttctccg caagaattga ttggctccaa ttcttggagt ggtgaatccg
     6301 ttagcgaggt gccgccggct tccattcagg tcgaggtggc ccggctccat gcaccgcgac
     6361 gcaacgcggg gaggcagaca aggtataggg cggcgcctac aatccatgcc aacccgttcc
     6421 atgtgctcgc cgaggcggca taaatcgccg tgacgatcag cggtccaatg atcgaagtta
     6481 ggctggtaag agccgcgagc gatccttgaa gctgtccctg atggtcgtca tctacctgcc
     6541 tggacagcat ggcctgcaac gcgggcatcc cgatgccgcc ggaagcgaga agaatcataa
     6601 tggggaaggc catccagcct cgcgtcgcga acgccagcaa gacgtagccc agcgcgtcgg
     6661 ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg tttggtggcg ggaccagtga
     6721 cgaaggcttg agcgagggcg tgcaagattc cgaataccgc aagcgacagg ccgatcatcg
     6781 tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc
     6841 ctacgagttg catgataaag aagacagtca taagtgcggc gacgatagtc atgccccgcg
     6901 cccaccggaa ggagctgact gggttgaagg ctctcaaggg catgggtcgg cgctctccct
     6961 tatgcgactc ctgcattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg
     7021 ccgcaaggaa tggtgcatgt aaggagatgg cgcccaacag tcccccggcc acggggcctg
     7081 ccaccatacc cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc cgatcttccc
     7141 catcggtgat gtcggcgata taggcgccag caaccgcacc tgtggcgccg gtgatgccgg
     7201 ccacgatgcg tccggcgtag agaatccaca ggacgggtgt ggtcgccatg atcgcgtagt
     7261 cgatagtggc tccaagtagc gaagcgagca ggactgggcg gcggccaaag cggtcggaca
     7321 gtgctccgag aacgggtgcg catagaaatt gcatcaacgc atatagcgct agcagcacgc
     7381 catagtgact ggcgatgctg tcggaatgga caatatcccg caagaggccc ggcagtaccg
     7441 gcataaccaa gcctatgcct acagcatcca gggtgacggt gccgaggatg acgatgagcg
     7501 cattgttaga tttcatacac ggtgcctgac tgcgttagca atttaactgt gataaactac
     7561 cgcattaaag ctaatcgatg ataagctgtc aaacatgaga attaattcat gttctttcct
     7621 gcgttatccc ctgattctgt ggataaccgt attaccgc
//
