LOCUS       40924_30795        8209 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pMGWA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8209)
  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 8209)
  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 8209)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8209)
  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..8209
                     /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             123..1220
                     /codon_start=1
                     /label=MBP
                     /note="maltose binding protein from E. coli"
                     /translation="KTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEE
                     KFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLI
                     AYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAA
                     DGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGET
                     AMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLE
                     NYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFW
                     YAVRTAVINAASGRQTVDEALKDAQT"
     protein_bind    1239..1363
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        1388..1418
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             1472..2128
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             2473..2775
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(2819..2943)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             2957..2974
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     terminator      3041..3088
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3125..3580
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        3607..3711
                     /label=AmpR promoter
     CDS             3712..4569
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      4743..5331
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(5517..5659)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(5764..5952)
                     /codon_start=1
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
                     /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     protein_bind    complement(6727..6748)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(6764..7843)
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(7844..7921)
                     /label=lacI promoter
ORIGIN
        1 agatccagat ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga
       61 taacaattcc cctctagaaa taattttgtt taactttaag aaggagatat accatgggta
      121 ccaaaactga agaaggtaaa ctggtaatct ggattaacgg cgataaaggc tataacggtc
      181 tcgctgaagt cggtaagaaa ttcgagaaag ataccggaat taaagtcacc gttgagcatc
      241 cggataaact ggaagagaaa ttcccacagg ttgcggcaac tggcgatggc cctgacatta
      301 tcttctgggc acacgaccgc tttggtggct acgctcaatc tggcctgttg gctgaaatca
      361 ccccggacaa agcgttccag gacaagctgt atccgtttac ctgggatgcc gtacgttaca
      421 acggcaagct gattgcttac ccgatcgctg ttgaagcgtt atcgctgatt tataacaaag
      481 atctgctgcc gaacccgcca aaaacctggg aagagatccc ggcgctggat aaagaactga
      541 aagcgaaagg taagagcgcg ctgatgttca acctgcaaga accgtacttc acctggccgc
      601 tgattgctgc tgacgggggt tatgcgttca agtatgaaaa cggcaagtac gacattaaag
      661 acgtgggcgt ggataacgct ggcgcgaaag cgggtctgac cttcctggtt gacctgatta
      721 aaaacaaaca catgaatgca gacaccgatt actccatcgc agaagctgcc tttaataaag
      781 gcgaaacagc gatgaccatc aacggcccgt gggcatggtc caacatcgac accagcaaag
      841 tgaattatgg tgtaacggta ctgccgacct tcaagggtca accatccaaa ccgttcgttg
      901 gcgtgctgag cgcaggtatt aacgccgcca gtccgaacaa agagctggca aaagagttcc
      961 tcgaaaacta tctgctgact gatgaaggtc tggaagcggt taataaagac aaaccgctgg
     1021 gtgccgtagc gctgaagtct tacgaggaag agttggcgaa agatccacgt attgccgcca
     1081 ccatggaaaa cgcccagaaa ggtgaaatca tgccgaacat cccgcagatg tccgctttct
     1141 ggtatgccgt gcgtactgcg gtgatcaacg ccgccagcgg tcgtcagact gtcgatgaag
     1201 ccctgaaaga cgcgcagact ggtaccggat cttacatcac aagtttgtac aaaaaagctg
     1261 aacgagaaac gtaaaatgat ataaatatca atatattaaa ttagattttg cataaaaaac
     1321 agactacata atactgtaaa acacaacata tccagtcact atggcggccg cattaggcac
     1381 cccaggcttt acactttatg cttccggctc gtataatgtg tggattttga gttaggatcc
     1441 gtcgagattt tcaggagcta aggaagctaa aatggagaaa aaaatcactg gatataccac
     1501 cgttgatata tcccaatggc atcgtaaaga acattttgag gcatttcagt cagttgctca
     1561 atgtacctat aaccagaccg ttcagctgga tattacggcc tttttaaaga ccgtaaagaa
     1621 aaataagcac aagttttatc cggcctttat tcacattctt gcccgcctga tgaatgctca
     1681 tccggaattc cgtatggcaa tgaaagacgg tgagctggtg atatgggata gtgttcaccc
     1741 ttgttacacc gttttccatg agcaaactga aacgttttca tcgctctgga gtgaatacca
     1801 cgacgatttc cggcagtttc tacacatata ttcgcaagat gtggcgtgtt acggtgaaaa
     1861 cctggcctat ttccctaaag ggtttattga gaatatgttt ttcgtctcag ccaatccctg
     1921 ggtgagtttc accagttttg atttaaacgt ggccaatatg gacaacttct tcgcccccgt
     1981 tttcaccatg ggcaaatatt atacgcaagg cgacaaggtg ctgatgccgc tggcgattca
     2041 ggttcatcat gccgtttgtg atggcttcca tgtcggcaga atgcttaatg aattacaaca
     2101 gtactgcgat gagtggcagg gcggggcgta aacgcgtgga tccggcttac taaaagccag
     2161 ataacagtat gcgtatttgc gcgctgattt ttgcggtata agaatatata ctgatatgta
     2221 tacccgaagt atgtcaaaaa gaggtatgct atgaagcagc gtattacagt gacagttgac
     2281 agcgacagct atcagttgct caaggcatat atgatgtcaa tatctccggt ctggtaagca
     2341 caaccatgca gaatgaagcc cgtcgtctgc gtgccgaacg ctggaaagcg gaaaatcagg
     2401 aagggatggc tgaggtcgcc cggtttattg aaatgaacgg ctcttttgct gacgagaaca
     2461 ggggctggtg aaatgcagtt taaggtttac acctataaaa gagagagccg ttatcgtctg
     2521 tttgtggatg tacagagtga tattattgac acgcccgggc gacggatggt gatccccctg
     2581 gccagtgcac gtctgctgtc agataaagtc tcccgtgaac tttacccggt ggtgcatatc
     2641 ggggatgaaa gctggcgcat gatgaccacc gatatggcca gtgtgccggt ctccgttatc
     2701 ggggaagaag tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc cattaacctg
     2761 atgttctggg gaatataaat gtcaggctcc cttatacaca gccagtctgc aggtcgacca
     2821 tagtgactgg atatgttgtg ttttacagta ttatgtagtc tgttttttat gcaaaatcta
     2881 atttaatata ttgatattta tatcatttta cgtttctcgt tcagctttct tgtacaaagt
     2941 ggtgatgtac ctcgagcacc accaccacca ccactgagat ccggctgcta acaaagcccg
     3001 aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc
     3061 ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg gattggcgaa
     3121 tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     3181 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     3241 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     3301 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     3361 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     3421 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     3481 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     3541 tttaacgcga attttaacaa aatattaacg tttacaattt caggtggcac ttttcgggga
     3601 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     3661 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     3721 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     3781 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     3841 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     3901 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     3961 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     4021 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     4081 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     4141 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     4201 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgcagca
     4261 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     4321 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     4381 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     4441 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     4501 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     4561 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     4621 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     4681 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     4741 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     4801 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     4861 ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac
     4921 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     4981 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     5041 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     5101 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     5161 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     5221 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     5281 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     5341 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     5401 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
     5461 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg
     5521 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatata tggtgcactc
     5581 tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg
     5641 tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     5701 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     5761 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc
     5821 gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt
     5881 ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc
     5941 ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc
     6001 gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact
     6061 ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac
     6121 tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca
     6181 gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag
     6241 actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt
     6301 gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag
     6361 gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg
     6421 ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt
     6481 gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat
     6541 cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg
     6601 tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg
     6661 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg
     6721 tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc
     6781 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     6841 gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc
     6901 ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca
     6961 ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat
     7021 cgtcgtatcc cactaccgag atatccgcac caacgcgcag cccggactcg gtaatggcgc
     7081 gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct
     7141 cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt
     7201 ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac
     7261 gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga
     7321 ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg
     7381 gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag
     7441 caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga
     7501 gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca
     7561 ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg
     7621 cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt
     7681 gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc
     7741 gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga
     7801 caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt
     7861 gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt
     7921 ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg
     7981 ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac
     8041 agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg
     8101 aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca
     8161 cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcg
//
