LOCUS       40924_18746        7553 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pEX18ApGW, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7553)
  AUTHORS   Choi KH, Schweizer HP.
  TITLE     An improved method for rapid generation of unmarked Pseudomonas 
            aeruginosa deletion mutants
  JOURNAL   BMC Microbiol. 5, 30 (2005)
  PUBMED    15907219
REFERENCE   2  (bases 1 to 7553)
  AUTHORS   Choi K-H., Schweizer HP.
  TITLE     Direct Submission
  JOURNAL   Submitted (11-FEB-2005) Microbiology, Immunology and Pathology, 
            Colorado State University, Fort Collins, CO 80523, USA
REFERENCE   3  (bases 1 to 7553)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7553)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "BMC 
            Microbiol. 5, 30 (2005)"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (11-FEB-2005) Microbiology, Immunology and Pathology, Colorado State
            University, Fort Collins, CO 80523, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7553
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(212..1630)
                     /codon_start=1
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     promoter        complement(1631..2076)
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     oriT            complement(2450..2559)
                     /direction=LEFT
                     /label=oriT
                     /note="incP origin of transfer"
     terminator      complement(2980..3007)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(3099..3185)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     primer_bind     3535..3551
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3596..3720
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(3764..4066)
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     CDS             complement(4411..5067)
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        complement(5121..5151)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     protein_bind    complement(5176..5300)
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(5332..5348)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(5356..5372)
                     /label=lac operator
                     /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(5380..5410)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(5425..5446)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(5734..6322)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6496..7353)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(7354..7458)
                     /label=AmpR promoter
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataatcg gcattttctt ttgcgttttt atttgttaac tgttaattgt ccttgttcaa
      241 ggatgctgtc tttgacaaca gatgttttct tgcctttgat gttcagcagg aagctaggcg
      301 caaacgttga ttgtttgtct gcgtagaatc ctctgtttgt catatagctt gtaatcacga
      361 cattgtttcc tttcgcttga ggtacagcga agtgtgagta agtaaaggtt acatcgttag
      421 gatcaagatc catttttaac acaaggccag ttttgttcag cggcttgtat gggccagtta
      481 aagaattaga aacataacca agcatgtaaa tatcgttaga cgtaatgccg tcaatcgtca
      541 tttttgatcc gcgggagtca gtgaacagat accatttgcc gttcatttta aagacgttcg
      601 cgcgttcaat ttcatctgtt actgtgttag atgcaatcag cggtttcatc acttttttca
      661 gtgtgtaatc atcgtttagc tcaatcatac cgagagcgcc gtttgctaac tcagccgtgc
      721 gttttttatc gctttgcaga agtttttgac tttcttgacg gaagaatgat gtgcttttgc
      781 catagtatgc tttgttaaat aaagattctt cgccttggta gccatcttca gttccagtgt
      841 ttgcttcaaa tactaagtat ttgtggcctt tatcttctac gtagtgagga tctctcagcg
      901 tatggttgtc gcctgagctg tagttgcctt catcgatgaa ctgctgtaca ttttgatacg
      961 tttttccgtc accgtcaaag attgatttat aatcctctac accgttgatg ttcaaagagc
     1021 tgtctgatgc tgatacgtta acttgtgcag ttgtcagtgt ttgtttgccg taatgtttac
     1081 cggagaaatc agtgtagaat aaacggattt ttccgtcaga tgtaaatgtg gctgaacctg
     1141 accattcttg tgtttggtct tttaggatag aatcatttgc atcgaatttg tcgctgtctt
     1201 taaagacgcg gccagcgttt ttccagctgt caatagaagt ttcgccgact ttttgataga
     1261 acatgtaaat cgatgtgtca tccgcatttt taggatctcc ggctaatgca aagacgatgt
     1321 ggtagccgtg atagtttgcg acagtgccgt cagcgttttg taatggccag ctgtcccaaa
     1381 cgtccaggcc ttttgcagaa gagatatttt taattgtgga cgaatcgaac tcaggaactt
     1441 gatatttttc atttttttgc tgttcaggga tttgcagcat atcatggcgt gtaatatggg
     1501 aaatgccgta tgtttcctta tatggctttt ggttcgtttc tttcgcaaac gcttgagttg
     1561 cgcctcctgc cagcagtgcg gtagtaaagg ttaatactgt tgcttgtttt gcaaactttt
     1621 tgatgttcat cgttcatgtc tcctttttta tgtactgtgt tagcggtctg cttcttccag
     1681 ccctcctgtt tgaagatggc aagttagtta cgcacaataa aaaaagacct aaaatatgta
     1741 aggggtgacg ccaaagtata cactttgccc tttacacatt ttaggtcttg cctgctttat
     1801 cagtaacaaa cccgcgcgat ttacttttcg acctcattct attagactct cgtttggatt
     1861 gcaactggtc tattttcctc ttttgtttga tagaaaatca taaaaggatt tgcagactac
     1921 gggcctaaag aactaaaaaa tctatctgtt tcttttcatt ctctgtattt tttatagttt
     1981 ctgttgcatg ggcataaagt tgccttttta atcacaattc agaaaatatc ataatatctc
     2041 atttcactaa ataatagtga acggcaggta tatgtgatgg gttaaaaagg atcgatcctc
     2101 tagctagagt cgatcttcgc cagcagggcg aggatcgtgg catcaccgaa ccgcgccgtg
     2161 cgcgggtcgt cggtgagcca gagtttcagc aggccgccca ggcggcccag gtcgccattg
     2221 atgcgggcca gctcgcggac gtgctcatag tccacgacgc ccgtgatttt gtagccctgg
     2281 ccgacggcca gcaggtaggc cgacaggctc atgccggccg ccgccgcctt ttcctcaatc
     2341 gctcttcgtt cgtctggaag gcagtacacc ttgataggtg ggctgccctt cctggttggc
     2401 ttggtttcat cagccatccg cttgccctca tctgttacgc cggcggtagc cggccagcct
     2461 cgcagagcag gattcccgtt gagcaccgcc aggtgcgaat aagggacagt gaagaaggaa
     2521 cacccgctcg cgggtgggcc tacttcacct atcctgcccg gctgacgccg ttggatacac
     2581 caaggaaagt ctacacgaac cctttggcaa aatcctgtat atcgtgcgaa aaaggatgga
     2641 tataccgaaa aaatcgctat aatgaccccg aagcagggtt atgcagcgga aaagcgctgc
     2701 ttccctgctg ttttgtggaa tatctaccga ctggaaacag gcaaatgcag gaaattactg
     2761 aactgagggg acaggcgaga gacgatgcca aagagctaca ccgacgagct ggccgagtgg
     2821 gttgaatccc gcgcggccaa gaagcgccgg cgtgatgagg ctgcggttgc gttcctggcg
     2881 gtgagggcgg atgtcgatat gcgtaaggag aaaataccgc atcaggcgca tatttgaatg
     2941 tatttagaaa aataaacaaa aagagtttgt agaaacgcaa aaaggccatc cgtcaggatg
     3001 gccttctgct taatttgatg cctggcagtt tatggcgggc gtcctgcccg ccaccctccg
     3061 ggccgttgct tcgcaacgtt caaatccgct cccggcggat ttgtcctact caggagagcg
     3121 ttcaccgaca aacaacagat aaaacgaaag gcccagtctt tcgactgagc ctttcgtttt
     3181 atttgatgcc tggcagttcc ctactctcgc atggggagac cccacactac catcggcgct
     3241 acggcgtttc acttctgagt tcggcatggg gtcaggtggg accaccgcgc tactgccgcc
     3301 aggcaaattc tgttttatca gaccgcttct gcgttctgat ttaatctgta tcaggctgaa
     3361 aatcttctct catccgccaa aacagccaag ctcgccattc gccattcagg ctgcgcaact
     3421 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
     3481 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
     3541 cgacggccag tgccaagctt gcatgcctgc aggtcgactc tagaggatcc ccatcaccac
     3601 tttgtacaag aaagctgaac gagaaacgta aaatgatata aatatcaata tattaaatta
     3661 gattttgcat aaaaaacaga ctacataata ctgtaaaaca caacatatcc agtcactatg
     3721 gtcgacctgc agactggctg tgtataacgg agcctgacat ttatattccc cagaacatca
     3781 ggttaatggc gtttttgatg tcattttcgc ggtggctgag atcagccact tcttccccga
     3841 taacggagac cggcacactg gccatatcgg tggtcatcat gcgccagctt tcatccccga
     3901 tatgcaccac cgggtaaagt tcacgggaga ctttatctga cagcagacgt gcactggcca
     3961 gggggatcac catccgtcgc ccgggcgtgt caataatatc actctgtaca tccacaaaca
     4021 gacgataacg gctctctctt ttataggtgt aaaccttaaa ctgcatttca ccagtccctg
     4081 ttctcgtcag caaaagagcc gttcatttca ataaaccggg cgacctcagc catcccttcc
     4141 tgattttccg ctttccagcg ttcggcacgc agacgacggg cttcattctg catggttgtg
     4201 cttaccagac cggagatatt gacatcatat atgccttgag caactgatag ctgtcgctgt
     4261 caactgtcac tgtaatacgc tgcttcatag cacacctctt tttgacatac ttcgggtata
     4321 catatcagta tatattctta taccgcaaaa atcagcgcgc aaatacgcat actgttatct
     4381 ggcttttagt aagccggatc cacgcgttta cgccccgccc tgccactcat cgcagtactg
     4441 ttgtaattca ttaagcattc tgccgacatg gaagccatca cagacggcat gatgaacctg
     4501 aatcgccagc ggcatcagca ccttgtcgcc ttgcgtataa tatttgccca tggtgaaaac
     4561 gggggcgaag aagttgtcca tattggccac gtttaaatca aaactggtga aactcaccca
     4621 gggattggct gagacgaaaa acatattctc aataaaccct ttagggaaat aggccaggtt
     4681 ttcaccgtaa cacgccacat cttgcgaata tatgtgtaga aactgccgga aatcgtcgtg
     4741 gtattcactc cagagcgatg aaaacgtttc agtttgctca tggaaaacgg tgtaacaagg
     4801 gtgaacacta tcccatatca ccagctcacc gtctttcatt gccatacgga attccggatg
     4861 agcattcatc aggcgggcaa gaatgtgaat aaaggccgga taaaacttgt gcttattttt
     4921 ctttacggtc tttaaaaagg ccgtaatatc cagctgaacg gtctggttat aggtacattg
     4981 agcaactgac tgaaatgcct caaaatgttc tttacgatgc cattgggata tatcaacggt
     5041 ggtatatcca gtgatttttt tctccatttt agcttcctta gctcctgaaa atctcgccgg
     5101 atcctaactc aaaatccaca cattatacga gccggaagca taaagtgtaa agcctggggt
     5161 gcctaatgcg gccgccatag tgactggata tgttgtgttt tacagtatta tgtagtctgt
     5221 tttttatgca aaatctaatt taatatattg atatttatat cattttacgt ttctcgttca
     5281 gcttttttgt acaaacttgt gatgggtacc gagctcgaat tcgtaatcat ggtcatagct
     5341 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     5401 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     5461 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     5521 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     5581 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     5641 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5701 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     5761 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     5821 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     5881 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     5941 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     6001 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     6061 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     6121 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     6181 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     6241 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6301 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6361 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     6421 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6481 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6541 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6601 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6661 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6721 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6781 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6841 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6901 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6961 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     7021 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     7081 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     7141 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     7201 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     7261 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     7321 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     7381 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     7441 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     7501 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtc
//
