LOCUS       40924_24543        7132 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pHGWA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7132)
  AUTHORS   Busso D, Delagoutte-Busso B, Moras D.
  TITLE     Construction of a set Gateway-based destination vectors for 
            high-throughput cloning and expression screening in Escherichia coli
  JOURNAL   Anal. Biochem. 343 (2), 313-321 (2005)
  PUBMED    15993367
REFERENCE   2  (bases 1 to 7132)
  AUTHORS   Busso D, Delagoutte-Busso B, Salim L, Moras D.
  TITLE     Direct Submission
  JOURNAL   Submitted (25-APR-2008) Structural Biology and Genomics Platform, 
            IGBMC, 1, rue Laurent Fries, Illkirch 67404, France
REFERENCE   3  (bases 1 to 7132)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7132)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Anal. 
            Biochem."; date: "2005"; volume: "343"; issue: "2"; pages: "313-321"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (25-APR-2008) Structural Biology and Genomics Platform, IGBMC, 1, 
            rue Laurent Fries, Illkirch 67404, France"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7132
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        27..45
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    46..70
                     /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)."
     RBS             85..107
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             126..143
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     protein_bind    162..286
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        311..341
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             395..1051
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             1396..1698
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(1742..1866)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             1880..1897
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     terminator      1964..2011
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2048..2503
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2530..2634
                     /label=AmpR promoter
     CDS             2635..3492
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      3666..4254
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(4440..4582)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(4687..4875)
                     /codon_start=1
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
                     /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     protein_bind    complement(5650..5671)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(5687..6766)
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(6767..6844)
                     /label=lacI promoter
ORIGIN
        1 agatccagat ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga
       61 taacaattcc cctctagaaa taattttgtt taactttaag aaggagatat accatgggca
      121 gcagccatca tcatcatcat cacggtaccg gatcttacat cacaagtttg tacaaaaaag
      181 ctgaacgaga aacgtaaaat gatataaata tcaatatatt aaattagatt ttgcataaaa
      241 aacagactac ataatactgt aaaacacaac atatccagtc actatggcgg ccgcattagg
      301 caccccaggc tttacacttt atgcttccgg ctcgtataat gtgtggattt tgagttagga
      361 tccgtcgaga ttttcaggag ctaaggaagc taaaatggag aaaaaaatca ctggatatac
      421 caccgttgat atatcccaat ggcatcgtaa agaacatttt gaggcatttc agtcagttgc
      481 tcaatgtacc tataaccaga ccgttcagct ggatattacg gcctttttaa agaccgtaaa
      541 gaaaaataag cacaagtttt atccggcctt tattcacatt cttgcccgcc tgatgaatgc
      601 tcatccggaa ttccgtatgg caatgaaaga cggtgagctg gtgatatggg atagtgttca
      661 cccttgttac accgttttcc atgagcaaac tgaaacgttt tcatcgctct ggagtgaata
      721 ccacgacgat ttccggcagt ttctacacat atattcgcaa gatgtggcgt gttacggtga
      781 aaacctggcc tatttcccta aagggtttat tgagaatatg tttttcgtct cagccaatcc
      841 ctgggtgagt ttcaccagtt ttgatttaaa cgtggccaat atggacaact tcttcgcccc
      901 cgttttcacc atgggcaaat attatacgca aggcgacaag gtgctgatgc cgctggcgat
      961 tcaggttcat catgccgttt gtgatggctt ccatgtcggc agaatgctta atgaattaca
     1021 acagtactgc gatgagtggc agggcggggc gtaaacgcgt ggatccggct tactaaaagc
     1081 cagataacag tatgcgtatt tgcgcgctga tttttgcggt ataagaatat atactgatat
     1141 gtatacccga agtatgtcaa aaagaggtat gctatgaagc agcgtattac agtgacagtt
     1201 gacagcgaca gctatcagtt gctcaaggca tatatgatgt caatatctcc ggtctggtaa
     1261 gcacaaccat gcagaatgaa gcccgtcgtc tgcgtgccga acgctggaaa gcggaaaatc
     1321 aggaagggat ggctgaggtc gcccggttta ttgaaatgaa cggctctttt gctgacgaga
     1381 acaggggctg gtgaaatgca gtttaaggtt tacacctata aaagagagag ccgttatcgt
     1441 ctgtttgtgg atgtacagag tgatattatt gacacgcccg ggcgacggat ggtgatcccc
     1501 ctggccagtg cacgtctgct gtcagataaa gtctcccgtg aactttaccc ggtggtgcat
     1561 atcggggatg aaagctggcg catgatgacc accgatatgg ccagtgtgcc ggtctccgtt
     1621 atcggggaag aagtggctga tctcagccac cgcgaaaatg acatcaaaaa cgccattaac
     1681 ctgatgttct ggggaatata aatgtcaggc tcccttatac acagccagtc tgcaggtcga
     1741 ccatagtgac tggatatgtt gtgttttaca gtattatgta gtctgttttt tatgcaaaat
     1801 ctaatttaat atattgatat ttatatcatt ttacgtttct cgttcagctt tcttgtacaa
     1861 agtggtgatg tacctcgagc accaccacca ccaccactga gatccggctg ctaacaaagc
     1921 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     1981 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc
     2041 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     2101 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     2161 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     2221 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     2281 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     2341 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     2401 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     2461 aaatttaacg cgaattttaa caaaatatta acgtttacaa tttcaggtgg cacttttcgg
     2521 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     2581 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     2641 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     2701 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     2761 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     2821 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     2881 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     2941 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     3001 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     3061 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     3121 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgca
     3181 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     3241 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     3301 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     3361 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     3421 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     3481 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     3541 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     3601 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     3661 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     3721 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     3781 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     3841 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     3901 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     3961 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     4021 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     4081 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     4141 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     4201 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     4261 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     4321 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     4381 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     4441 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat atatggtgca
     4501 ctctcagtac aatctgctct gatgccgcat agttaagcca gtatacactc cgctatcgct
     4561 acgtgactgg gtcatggctg cgccccgaca cccgccaaca cccgctgacg cgccctgacg
     4621 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     4681 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt aaagctcatc
     4741 agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca gctcgttgag
     4801 tttctccaga agcgttaatg tctggcttct gataaagcgg gccatgttaa gggcggtttt
     4861 ttcctgtttg gtcactgatg cctccgtgta agggggattt ctgttcatgg gggtaatgat
     4921 accgatgaaa cgagagagga tgctcacgat acgggttact gatgatgaac atgcccggtt
     4981 actggaacgt tgtgagggta aacaactggc ggtatggatg cggcgggacc agagaaaaat
     5041 cactcagggt caatgccagc gcttcgttaa tacagatgta ggtgttccac agggtagcca
     5101 gcagcatcct gcgatgcaga tccggaacat aatggtgcag ggcgctgact tccgcgtttc
     5161 cagactttac gaaacacgga aaccgaagac cattcatgtt gttgctcagg tcgcagacgt
     5221 tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt gattcattct gctaaccagt
     5281 aaggcaaccc cgccagccta gccgggtcct caacgacagg agcacgatca tgcgcacccg
     5341 tggggccgcc atgccggcga taatggcctg cttctcgccg aaacgtttgg tggcgggacc
     5401 agtgacgaag gcttgagcga gggcgtgcaa gattccgaat accgcaagcg acaggccgat
     5461 catcgtcgcg ctccagcgaa agcggtcctc gccgaaaatg acccagagcg ctgccggcac
     5521 ctgtcctacg agttgcatga taaagaagac agtcataagt gcggcgacga tagtcatgcc
     5581 ccgcgcccac cggaaggagc tgactgggtt gaaggctctc aagggcatcg gtcgagatcc
     5641 cggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca
     5701 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     5761 tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat
     5821 tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca
     5881 gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag ctgtcttcgg
     5941 tatcgtcgta tcccactacc gagatatccg caccaacgcg cagcccggac tcggtaatgg
     6001 cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc
     6061 cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc
     6121 gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca
     6181 gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg
     6241 cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata atactgttga
     6301 tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca
     6361 cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg
     6421 cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca
     6481 ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg
     6541 gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca
     6601 gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt
     6661 cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag
     6721 agacaccggc atactctgcg acatcgtata acgttactgg tttcacattc accaccctga
     6781 attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg
     6841 tgtccgggat ctcgacgctc tcccttatgc gactcctgca ttaggaagca gcccagtagt
     6901 aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga gatggcgccc
     6961 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
     7021 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
     7081 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat cg
//
