LOCUS       40924_22789        7947 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pGTVL2, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7947)
  AUTHORS   Savitsky P, Burgess-Brown N, Gileadi O.
  TITLE     Direct Submission
  JOURNAL   Submitted (09-MAR-2011) Structural Genomics Consortium, University 
            of Oxford, Old Road Campus Research Building, Roosevelt Drive, 
            Oxford, Oxfordshire OX3 7DQ, UK
REFERENCE   2  (bases 1 to 7947)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 7947)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Submitted 
            (09-MAR-2011) Structural Genomics Consortium, University of Oxford, 
            Old Road Campus Research Building, Roosevelt Drive, Oxford, 
            Oxfordshire OX3 7DQ, UK"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7947
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             12..34
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             45..62
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     CDS             69..722
                     /codon_start=1
                     /label=GST
                     /note="glutathione S-transferase from Schistosoma
                     japonicum"
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             750..770
                     /codon_start=1
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
                     /translation="ENLYFQS"
     promoter        786..1231
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             1232..2650
                     /codon_start=1
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYKVPEFDSSTIKNISSAKGLDVWDSWPLQNTDGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     misc_feature    2699..2713
                     /label=3'-LIC
                     /note="3'-LIC"
     CDS             2761..2778
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     terminator      2845..2892
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2929..3384
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     CDS             complement(3480..4292)
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      4414..5002
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(5188..5330)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(5435..5623)
                     /codon_start=1
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
                     /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     protein_bind    complement(6398..6419)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(6435..7514)
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(7515..7592)
                     /label=lacI promoter
     promoter        7901..7919
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    7920..7944
                     /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)."
ORIGIN
        1 ctagaaataa ttttgtttaa ctttaagaag gagatataca tatgcaccat catcatcatc
       61 attcttctat gtcccctata ctaggttatt ggaaaattaa gggccttgtg caacccactc
      121 gacttctttt ggaatatctt gaagaaaaat atgaagagca tttgtatgag cgcgatgaag
      181 gtgataaatg gcgaaacaaa aagtttgaat tgggtttgga gtttcccaat cttccttatt
      241 atattgatgg tgatgttaaa ttaacacagt ctatggccat catacgttat atagctgaca
      301 agcacaacat gttgggtggt tgtccaaaag agcgtgcaga gatttcaatg cttgaaggag
      361 cggttttgga tattagatac ggtgtttcga gaattgcata tagtaaagac tttgaaactc
      421 tcaaagttga ttttcttagc aagctacctg aaatgctgaa aatgttcgaa gatcgtttat
      481 gtcataaaac atatttaaat ggtgatcatg taacccatcc tgacttcatg ttgtatgacg
      541 ctcttgatgt tgttttatac atggacccaa tgtgcctgga tgcgttccca aaattagttt
      601 gttttaaaaa acgtattgaa gctatcccac aaattgataa gtacttgaaa tccagcaagt
      661 atatagcatg gcctttgcag ggctggcaag ccacgtttgg tggtggcgac catcctccaa
      721 aatcgagctc aggtgtagat ctgggtaccg agaacctgta cttccaatcc atggagaccg
      781 acgtccacat atacctgccg ttcactatta tttagtgaaa tgagatatta tgatattttc
      841 tgaattgtga ttaaaaaggc aactttatgc ccatgcaaca gaaactataa aaaatacaga
      901 gaatgaaaag aaacagatag attttttagt tctttaggcc cgtagtctgc aaatcctttt
      961 atgattttct atcaaacaaa agaggaaaat agaccagttg caatccaaac gagagtctaa
     1021 tagaatgagg tcgaaaagta aatcgcgcgg gtttgttact gataaagcag gcaagaccta
     1081 aaatgtgtaa agggcaaagt gtatactttg gcgtcacccc ttacatattt taggtctttt
     1141 tttattgtgc gtaactaact tgccatcttc aaacaggagg gctggaagaa gcagaccgct
     1201 aacacagtac ataaaaaagg agacatgaac gatgaacatc aaaaagtttg caaaacaagc
     1261 aacagtatta acctttacta ccgcactgct ggcaggaggc gcaactcaag cgtttgcgaa
     1321 agaaacgaac caaaagccat ataaggaaac atacggcatt tcccatatta cacgccatga
     1381 tatgctgcaa atccctgaac agcaaaaaaa tgaaaaatat aaagttcctg agttcgattc
     1441 gtccacaatt aaaaatatct cttctgcaaa aggcctggac gtttgggaca gctggccatt
     1501 acaaaacact gacggcactg tcgcaaacta tcacggctac cacatcgtct ttgcattagc
     1561 cggagatcct aaaaatgcgg atgacacatc gatttacatg ttctatcaaa aagtcggcga
     1621 aacttctatt gacagctgga aaaacgctgg ccgcgtcttt aaagacagcg acaaattcga
     1681 tgcaaatgat tctatcctaa aagaccaaac acaagaatgg tcaggttcag ccacatttac
     1741 atctgacgga aaaatccgtt tattctacac tgatttctcc ggtaaacatt acggcaaaca
     1801 aacactgaca actgcacaag ttaacgtatc agcatcagac agctctttga acatcaacgg
     1861 tgtagaggat tataaatcaa tctttgacgg tgacggaaaa acgtatcaaa atgtacagca
     1921 gttcatcgat gaaggcaact acagctcagg cgacaaccat acgctgagag atcctcacta
     1981 cgtagaagat aaaggccaca aatacttagt atttgaagca aacactggaa ctgaagatgg
     2041 ctaccaaggc gaagaatctt tatttaacaa agcatactat ggcaaaagca catcattctt
     2101 ccgtcaagaa agtcaaaaac ttctgcaaag cgataaaaaa cgcacggctg agttagcaaa
     2161 cggcgctctc ggtatgattg agctaaacga tgattacaca ctgaaaaaag tgatgaaacc
     2221 gctgattgca tctaacacag taacagatga aattgaacgc gcgaacgtct ttaaaatgaa
     2281 cggcaaatgg tacctgttca ctgactcccg cggatcaaaa atgacgattg acggcattac
     2341 gtctaacgat atttacatgc ttggttatgt ttctaattct ttaactggcc catacaagcc
     2401 gctgaacaaa actggccttg tgttaaaaat ggatcttgat cctaacgatg taacctttac
     2461 ttactcacac ttcgctgtac ctcaagcgaa aggaaacaat gtcgtgatta caagctatat
     2521 gacaaacaga ggattctacg cagacaaaca atcaacgttt gcgcctagct tcctgctgaa
     2581 catcaaaggc aagaaaacat ctgttgtcaa agacagcatc cttgaacaag gacaattaac
     2641 agttaacaaa taaaaacgca aaagaaaatg ccgatatcct attggcattg acggtctcca
     2701 gtaaaggtgg atacggatcc gaattcgagc tccgtcgaca agcttgcggc cgcactcgag
     2761 caccaccacc accaccactg agatccggct gctaacaaag cccgaaagga agctgagttg
     2821 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg
     2881 aggggttttt tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta
     2941 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     3001 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     3061 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     3121 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     3181 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     3241 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     3301 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     3361 acaaaatatt aacgtttaca atttcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     3421 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaat taattcttag
     3481 aaaaactcat cgagcatcaa atgaaactgc aatttattca tatcaggatt atcaatacca
     3541 tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca gttccatagg
     3601 atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat acaacctatt
     3661 aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt gacgactgaa
     3721 tccggtgaga atggcaaaag tttatgcatt tctttccaga cttgttcaac aggccagcca
     3781 ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg tgattgcgcc
     3841 tgagcgagac gaaatacgcg atcgctgtta aaaggacaat tacaaacagg aatcgaatgc
     3901 aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc aggatattct
     3961 tctaatacct ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca tgcatcatca
     4021 ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag ccagtttagt
     4081 ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt cagaaacaac
     4141 tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg cccgacatta
     4201 tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa tcgcggccta
     4261 gagcaagacg tttcccgttg aatatggctc ataacacccc ttgtattact gtttatgtaa
     4321 gcagacagtt ttattgttca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     4381 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     4441 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     4501 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     4561 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     4621 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     4681 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     4741 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     4801 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     4861 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     4921 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     4981 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     5041 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     5101 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     5161 gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg
     5221 cggtatttca caccgcatat atggtgcact ctcagtacaa tctgctctga tgccgcatag
     5281 ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc
     5341 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     5401 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     5461 gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg aagcgattca cagatgtctg
     5521 cctgttcatc cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga
     5581 taaagcgggc catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaag
     5641 ggggatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac
     5701 gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg
     5761 tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcgc ttcgttaata
     5821 cagatgtagg tgttccacag ggtagccagc agcatcctgc gatgcagatc cggaacataa
     5881 tggtgcaggg cgctgacttc cgcgtttcca gactttacga aacacggaaa ccgaagacca
     5941 ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc
     6001 gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc cgggtcctca
     6061 acgacaggag cacgatcatg cgcacccgtg gggccgccat gccggcgata atggcctgct
     6121 tctcgccgaa acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga
     6181 ttccgaatac cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag cggtcctcgc
     6241 cgaaaatgac ccagagcgct gccggcacct gtcctacgag ttgcatgata aagaagacag
     6301 tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg actgggttga
     6361 aggctctcaa gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa
     6421 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     6481 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt
     6541 ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc
     6601 agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac
     6661 ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatatccgca
     6721 ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg
     6781 gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa
     6841 ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg
     6901 agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct
     6961 aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg
     7021 tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac
     7081 gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag
     7141 ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct
     7201 tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga
     7261 gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg
     7321 ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc
     7381 agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg
     7441 ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac
     7501 gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata
     7561 ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga
     7621 ctcctgcatt aggaagcagc ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag
     7681 gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat
     7741 acccacgccg aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt
     7801 gatgtcggcg atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat
     7861 gcgtccggcg tagaggatcg agatctcgat cccgcgaaat taatacgact cactataggg
     7921 gaattgtgag cggataacaa ttcccct
//
