LOCUS       40924_24737        8236 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pHMGWA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8236)
  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 8236)
  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 8236)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8236)
  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..8236
                     /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"
     CDS             150..1247
                     /codon_start=1
                     /label=MBP
                     /note="maltose binding protein from E. coli"
                     /translation="KTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEE
                     KFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLI
                     AYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAA
                     DGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGET
                     AMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLE
                     NYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFW
                     YAVRTAVINAASGRQTVDEALKDAQT"
     protein_bind    1266..1390
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        1415..1445
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             1499..2155
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             2500..2802
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(2846..2970)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             2984..3001
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     terminator      3068..3115
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3152..3607
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        3634..3738
                     /label=AmpR promoter
     CDS             3739..4596
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      4770..5358
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(5544..5686)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(5791..5979)
                     /codon_start=1
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
                     /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     protein_bind    complement(6754..6775)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(6791..7870)
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(7871..7948)
                     /label=lacI promoter
ORIGIN
        1 agatccagat ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga
       61 taacaattcc cctctagaaa taattttgtt taactttaag aaggagatat accatgggca
      121 gcagccatca tcatcatcat cacggtacca aaactgaaga aggtaaactg gtaatctgga
      181 ttaacggcga taaaggctat aacggtctcg ctgaagtcgg taagaaattc gagaaagata
      241 ccggaattaa agtcaccgtt gagcatccgg ataaactgga agagaaattc ccacaggttg
      301 cggcaactgg cgatggccct gacattatct tctgggcaca cgaccgcttt ggtggctacg
      361 ctcaatctgg cctgttggct gaaatcaccc cggacaaagc gttccaggac aagctgtatc
      421 cgtttacctg ggatgccgta cgttacaacg gcaagctgat tgcttacccg atcgctgttg
      481 aagcgttatc gctgatttat aacaaagatc tgctgccgaa cccgccaaaa acctgggaag
      541 agatcccggc gctggataaa gaactgaaag cgaaaggtaa gagcgcgctg atgttcaacc
      601 tgcaagaacc gtacttcacc tggccgctga ttgctgctga cgggggttat gcgttcaagt
      661 atgaaaacgg caagtacgac attaaagacg tgggcgtgga taacgctggc gcgaaagcgg
      721 gtctgacctt cctggttgac ctgattaaaa acaaacacat gaatgcagac accgattact
      781 ccatcgcaga agctgccttt aataaaggcg aaacagcgat gaccatcaac ggcccgtggg
      841 catggtccaa catcgacacc agcaaagtga attatggtgt aacggtactg ccgaccttca
      901 agggtcaacc atccaaaccg ttcgttggcg tgctgagcgc aggtattaac gccgccagtc
      961 cgaacaaaga gctggcaaaa gagttcctcg aaaactatct gctgactgat gaaggtctgg
     1021 aagcggttaa taaagacaaa ccgctgggtg ccgtagcgct gaagtcttac gaggaagagt
     1081 tggcgaaaga tccacgtatt gccgccacca tggaaaacgc ccagaaaggt gaaatcatgc
     1141 cgaacatccc gcagatgtcc gctttctggt atgccgtgcg tactgcggtg atcaacgccg
     1201 ccagcggtcg tcagactgtc gatgaagccc tgaaagacgc gcagactggt accggatctt
     1261 acatcacaag tttgtacaaa aaagctgaac gagaaacgta aaatgatata aatatcaata
     1321 tattaaatta gattttgcat aaaaaacaga ctacataata ctgtaaaaca caacatatcc
     1381 agtcactatg gcggccgcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     1441 taatgtgtgg attttgagtt aggatccgtc gagattttca ggagctaagg aagctaaaat
     1501 ggagaaaaaa atcactggat ataccaccgt tgatatatcc caatggcatc gtaaagaaca
     1561 ttttgaggca tttcagtcag ttgctcaatg tacctataac cagaccgttc agctggatat
     1621 tacggccttt ttaaagaccg taaagaaaaa taagcacaag ttttatccgg cctttattca
     1681 cattcttgcc cgcctgatga atgctcatcc ggaattccgt atggcaatga aagacggtga
     1741 gctggtgata tgggatagtg ttcacccttg ttacaccgtt ttccatgagc aaactgaaac
     1801 gttttcatcg ctctggagtg aataccacga cgatttccgg cagtttctac acatatattc
     1861 gcaagatgtg gcgtgttacg gtgaaaacct ggcctatttc cctaaagggt ttattgagaa
     1921 tatgtttttc gtctcagcca atccctgggt gagtttcacc agttttgatt taaacgtggc
     1981 caatatggac aacttcttcg cccccgtttt caccatgggc aaatattata cgcaaggcga
     2041 caaggtgctg atgccgctgg cgattcaggt tcatcatgcc gtttgtgatg gcttccatgt
     2101 cggcagaatg cttaatgaat tacaacagta ctgcgatgag tggcagggcg gggcgtaaac
     2161 gcgtggatcc ggcttactaa aagccagata acagtatgcg tatttgcgcg ctgatttttg
     2221 cggtataaga atatatactg atatgtatac ccgaagtatg tcaaaaagag gtatgctatg
     2281 aagcagcgta ttacagtgac agttgacagc gacagctatc agttgctcaa ggcatatatg
     2341 atgtcaatat ctccggtctg gtaagcacaa ccatgcagaa tgaagcccgt cgtctgcgtg
     2401 ccgaacgctg gaaagcggaa aatcaggaag ggatggctga ggtcgcccgg tttattgaaa
     2461 tgaacggctc ttttgctgac gagaacaggg gctggtgaaa tgcagtttaa ggtttacacc
     2521 tataaaagag agagccgtta tcgtctgttt gtggatgtac agagtgatat tattgacacg
     2581 cccgggcgac ggatggtgat ccccctggcc agtgcacgtc tgctgtcaga taaagtctcc
     2641 cgtgaacttt acccggtggt gcatatcggg gatgaaagct ggcgcatgat gaccaccgat
     2701 atggccagtg tgccggtctc cgttatcggg gaagaagtgg ctgatctcag ccaccgcgaa
     2761 aatgacatca aaaacgccat taacctgatg ttctggggaa tataaatgtc aggctccctt
     2821 atacacagcc agtctgcagg tcgaccatag tgactggata tgttgtgttt tacagtatta
     2881 tgtagtctgt tttttatgca aaatctaatt taatatattg atatttatat cattttacgt
     2941 ttctcgttca gctttcttgt acaaagtggt gatgtacctc gagcaccacc accaccacca
     3001 ctgagatccg gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga
     3061 gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa
     3121 aggaggaact atatccggat tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg
     3181 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     3241 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     3301 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     3361 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     3421 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     3481 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     3541 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt
     3601 acaatttcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     3661 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     3721 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     3781 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     3841 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     3901 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     3961 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     4021 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     4081 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     4141 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     4201 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     4261 gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact attaactggc
     4321 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     4381 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     4441 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     4501 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     4561 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     4621 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     4681 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     4741 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     4801 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     4861 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     4921 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     4981 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     5041 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     5101 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     5161 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     5221 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     5281 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     5341 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     5401 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     5461 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     5521 gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt
     5581 atttcacacc gcatatatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     5641 gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc
     5701 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     5761 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     5821 gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg
     5881 ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa
     5941 gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg
     6001 atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt
     6061 tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg
     6121 gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga
     6181 tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt
     6241 gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca
     6301 tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat
     6361 cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga
     6421 caggagcacg atcatgcgca cccgtggggc cgccatgccg gcgataatgg cctgcttctc
     6481 gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt gcaagattcc
     6541 gaataccgca agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa
     6601 aatgacccag agcgctgccg gcacctgtcc tacgagttgc atgataaaga agacagtcat
     6661 aagtgcggcg acgatagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc
     6721 tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc
     6781 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     6841 cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca
     6901 ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca
     6961 agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg
     7021 ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa
     7081 cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa
     7141 ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg
     7201 acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat
     7261 atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca
     7321 gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt
     7381 catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg
     7441 gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa
     7501 tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga
     7561 cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt
     7621 taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa
     7681 tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct
     7741 ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca
     7801 ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta
     7861 ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc
     7921 gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc
     7981 tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat
     8041 ggtgcatgca aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc
     8101 acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg
     8161 tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt
     8221 ccggcgtaga ggatcg
//
