LOCUS       40924_17621        9763 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pESP-4, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9763)
  AUTHORS   Grafsky AJ, Lu Q.
  TITLE     Direct Submission
  JOURNAL   Submitted (05-FEB-1999) Technical Services, Stratagene, 11011 N. 
            Torrey Pines Rd., La Jolla, CA 92037, USA
REFERENCE   2  (bases 1 to 9763)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 9763)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Submitted 
            (05-FEB-1999) Technical Services, Stratagene, 11011 N. Torrey Pines 
            Rd., La Jolla, CA 92037, USA"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..9763
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(138..921)
                     /direction=LEFT
                     /label=ars1
                     /note="Schizosaccharomyces pombe autonomously replicating 
                     sequence ars1"
     promoter        1296..1400
                     /label=AmpR promoter
     CDS             1401..2258
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      2432..3020
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    3308..3329
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        3344..3374
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3382..3398
                     /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)."
     primer_bind     3406..3422
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3443..3461
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     3491..3507
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(3915..5006)
                     /codon_start=1
                     /label=LEU2
                     /note="3-isopropylmalate dehydrogenase, required for
                     leucine biosynthesis"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVDPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        5962..7119
                     /label=nmt1 promoter
                     /note="wild-type nmt1 promoter from Schizosaccharomyces
                     pombe, conferring strong thiamine-repressible expression"
     CDS             7121..7774
                     /codon_start=1
                     /label=GST
                     /note="glutathione S-transferase from Schistosoma
                     japonicum"
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             7781..7798
                     /codon_start=1
                     /label=thrombin site
                     /note="thrombin recognition and cleavage site"
                     /translation="LVPRGS"
     CDS             7799..7822
                     /codon_start=1
                     /label=FLAG
                     /note="FLAG(R) epitope tag, followed by an enterokinase
                     cleavage site"
                     /translation="DYKDDDDK"
     promoter        complement(8886..8904)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(8914..8930)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      9072..9527
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagaattc gagtctaact ccttaaccac tcggacatag tgacttatct gacactcatt
      121 gtaaattaat atatatatag gcattttgtt tagttaaagg tacttaagta attagtataa
      181 acgaaccaat tttataatca ggaagttaag tgaatggtag cacatgtcgt aaaaattgtg
      241 aatttttatt gaataatatt ttaaatacaa gcctttctag actaggtata ctcataaaca
      301 tatatgagca aaaggataga ggagattaca ttgcatcttc tacaaattta tttattgccc
      361 tttactgaaa aattaaataa tgagtactaa atgataaaaa gcgctcagta caagaaagat
      421 tgcaaaaata ttgcattctt catgaattaa agttgcatat aaggcatatt gaaagtaata
      481 gtactaaaac agcagttagc gaaaattaat agaattatat tcgcaagaca attgtgacat
      541 taaattaaaa aattgtagaa tttttactat cctctttaac gccatgagcc tttataaaaa
      601 ggttaaatta gttttaacat tctttttttg agtaagagtt tacaatttat caaaacctgt
      661 gttattatat tcattagttt caatttatta gcatctagag aaaaatcaat tggcagttac
      721 attgttggaa tttatgaaga aaaagaatct acaacggaga ataagttgct gatcgctttc
      781 cctaaaattg tatattttgc tgagcttatt ttgacatttc gttgaagttt tctctaattc
      841 gcattcattt taagtaaaac aatgagaaat aaaattacaa aaaatacaaa ttaaaataca
      901 atttttagct ataattatag acgatgccct tgtatcccat tctgtctcgc ttgcccctac
      961 tttttatctt ttatatacca taatgaacgc tgccgctact aaccataccc cgattttaca
     1021 tttcggactc ccaaggacgt acaaaataga aaactataga aaaaaataat cagaaaatag
     1081 catgtcatct ctttgtaaaa cgcgtttgca agaagaaagg aaacaatgga gaagagatca
     1141 tccatttgta tgtaaatttt agtaaacttg aagaaatcac taacaacttc tcttacttag
     1201 ggattctatg caaaaccttg taaatcatct gatggaggac tcgatttaat gaattggaag
     1261 gttggaattc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     1321 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     1381 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     1441 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     1501 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     1561 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     1621 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     1681 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     1741 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     1801 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     1861 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     1921 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     1981 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     2041 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     2101 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     2161 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     2221 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     2281 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     2341 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     2401 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     2461 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     2521 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     2581 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     2641 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     2701 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     2761 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     2821 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     2881 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     2941 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     3001 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     3061 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     3121 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     3181 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     3241 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     3301 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     3361 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     3421 accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctgggtaccg
     3481 ggccccccct cgaggtcgac ggtatcgata agcttactga tgatgatgtg tctactttca
     3541 ggcgggtctg tagtaaggag aatgacatta taaagctggc acttagaatt ccacggacta
     3601 tagactatac tagtatactc cgtctactgt acgatacact tccgctcagg tccttgtcct
     3661 ttaacgaggc cttaccactc ttttgttact ctattgatcc agctcagcaa aggcagtgtg
     3721 atctaagatt ctatcttcgc gatgtagtaa aactagctag accgagaaag agactagaaa
     3781 tgcaaaaggc acttctacaa tggctgccat cattattatc cgatgtgacg ctgcattttt
     3841 tttttttttt tttttttttt tttttttttt tttttttttt tgtacaaata tcataaaaaa
     3901 agagaatctt tttaagcaag gattttctta acttcttcgg cgacagcatc accgacttcg
     3961 gtggtactgt tggaaccacc taaatcacca gttctgatac ctgcatccaa aaccttttta
     4021 actgcatctt caatggcttt accttcttca ggcaagttca atgacaattt caacatcatt
     4081 gcagcagaca agatagtggc gatagggtcg accttattct ttggcaaatc tggagcggaa
     4141 ccatggcatg gttcgtacaa accaaatgcg gtgttcttgt ctggcaaaga ggccaaggac
     4201 gcagatggca acaaacccaa ggaacctggg ataacggagg cttcatcgga gatgatatca
     4261 ccaaacatgt tgctggtgat tataatacca tttaggtggg ttgggttctt aactaggatc
     4321 atggcggcag aatcaatcaa ttgatgttga accttcaatg tagggaattc gttcttgatg
     4381 gtttcctcca cagtttttct ccataatctt gaagaggcca aaacattagc tttatccaag
     4441 gaccaaatag gcaatggtgg ctcatgttgt agggccatga aagcggccat tcttgtgatt
     4501 ctttgcactt ctggaacggt gtattgttca ctatcccaag cgacaccatc accatcgtct
     4561 tcctttctct taccaaagta aatacctccc actaattctc tgacaacaac gaagtcagta
     4621 cctttagcaa attgtggctt gattggagat aagtctaaaa gagagtcgga tgcaaagtta
     4681 catggtctta agttggcgta caattgaagt tctttacgga tttttagtaa accttgttca
     4741 ggtctaacac taccggtacc ccatttagga ccacccacag cacctaacaa aacggcatca
     4801 accttcttgg aggcttccag cgcctcatct ggaagtggga cacctgtagc atcgatagca
     4861 gcaccaccaa ttaaatgatt ttcgaaatcg aacttgacat tggaacgaac atcagaaata
     4921 gctttaagaa ccttaatggc ttcggctgtg atttcttgac caacgtggtc acctggcaaa
     4981 acgacgatct tcttaggggc agacattaga atggtatatc cttgaaatat atataaaaaa
     5041 aaaaaaaaaa aaaaaaaaaa aaaatgcagc ttctcaatga tattcgaata cgctttgagg
     5101 agatacagcc taatatccga caaactgttt tacagattta cgatcgtact tgttacccat
     5161 cattgaattt tgaacatccg aacctgggag ttttccctga aacagatagt atatttgaac
     5221 ctgtataata atatatagtc tagcgcttta cggaagacaa tgtatgtatt tcggttcctg
     5281 gagaaactat tgcatctatt gcataggtaa tcttgcacgt cgcatccccg gttcattttc
     5341 tgcgtttcca tcttgcactt caatagcata tctttgttaa cgaagcatct gtgcttcatt
     5401 ttgtagaaca aaaatgcaac gcgagagcgc taatttttca aacaaagaat ctgagctgca
     5461 tttttacaga acagaaatgc aacgcgaaag cgctatttta ccaacgaaga atctgtgctt
     5521 catttttgta aaacaaaaat gcaacgcgag agcgctaatt tttcaaacaa agaatctgag
     5581 ctgcattttt acagaacaga aatgcaacgc gagagcgcta ttttaccaac aaagaatcta
     5641 tacttctttt ttgttctaca aaaatgcaca cgtagcctgc atagtaccga gaaactagtg
     5701 cgaagtagtg atcaggtatt gctgttatct gatgagtata cgttgtcctg gccacggcag
     5761 aagcacgctt atcgctccaa tttcccacaa cattagtcaa ctccgttagg cccttcattg
     5821 aaagaaatga ggtcatcaaa tgtcttccaa tgtgagattt tgggccattt tttatagcaa
     5881 agattgaata aggcgcattt ttcttcaaag cttgatatcg aattcctgca ggtcgatcga
     5941 ctctagagga tcagaaaatt atcgccataa aagacagaat aagtcatcag cggttgtttc
     6001 atttcctata tttttttttt atttttttat tttttaataa gggaaaattt aacgtctaag
     6061 gatacagaag attgttagca cattaaagta ataaaggctt aagtagtaag tgccttagca
     6121 tgttattgta tttcaaagga cataatctaa aataataaca atatcatttc tcacaagtta
     6181 ttcaattttc tttttttttt ctaataatat caagaatgta ttatttgttt gacataagtc
     6241 aactaattta tttaatatgc tggattaatc ttgcagacat gtaaattaac aagttttagt
     6301 caaataacgt tgaagtttca atgaactcaa ataatttctc ttttttttta tataaccata
     6361 atctgattta tattttccgc aggatcaact gaagttatga catttggatt ggatcactta
     6421 taaccttggt cgccaaataa tacaaaaatc agcgttataa aacaaagaag gtttttgtta
     6481 agaaattaat cctctttctt gataagaaag ttgaaccgaa attgcagata ctgatatatg
     6541 aaaataatac ccacaatttt gggaatagcg caagcctcaa tttaaacaat aggtgaggac
     6601 acatgataat gacctcaatg attgttagaa gaaaagagcc tcattacaaa atcgaaaaat
     6661 gaatggttgg gtacaagttt ccaaaacatg gtaaagtgga ctttgcgtat gagacgtaaa
     6721 tagaaaaaaa cacttgttat atgttttcta gaattattgt tgtctcttta tggttggatg
     6781 atgcaaaata gtaatttcgg ttagttgctg taaaacacca cgagacaaat agatatggat
     6841 atttattaaa tcaggaaaaa cgtaactctc ggctactgga tggttcagtc acccaacgat
     6901 tactggggag agaaaacagg gcaaaagcaa agcttaaagg aatccgattg tcattcggca
     6961 atgtgcagcg aaactaaaaa ccggataatg gacctgttaa tcgaaacatt gaagatatat
     7021 aaaggaagag gaatcctggc atatcatcaa ttgaataagt tgaattaatt atttcaatct
     7081 cattctcact ttctgactta tagtcgcttt gttaaatcat atgtccccta tactaggtta
     7141 ttggaaaatt aagggccttg tgcaacccac tcgacttctt ttggaatatc ttgaagaaaa
     7201 atatgaagag catttgtatg agcgcgatga aggtgataaa tggcgaaaca aaaagtttga
     7261 attgggtttg gagtttccca atcttcctta ttatattgat ggtgatgtta aattaacaca
     7321 gtctatggcc atcatacgtt atatagctga caagcacaac atgttgggtg gttgtccaaa
     7381 agagcgtgca gagatttcaa tgcttgaagg agcggttttg gatattagat acggtgtttc
     7441 gagaattgca tatagtaaag actttgaaac tctcaaagtt gattttctta gcaagctacc
     7501 tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa acatatttaa atggtgatca
     7561 tgtaacccat cctgacttca tgttgtatga cgctcttgat gttgttttat acatggaccc
     7621 aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa aaacgtattg aagctatccc
     7681 acaaattgat aagtacttga aatccagcaa gtatatagca tggcctttgc agggctggca
     7741 agccacgttt ggtggtggcg accatcctcc aaaatcggat ctggttccgc gtggatccga
     7801 ctacaaggat gacgacgaca agcatgcatc tgctagcccg ggccatggag atctgcatgc
     7861 gggtcttgtt ccatgagggt aaaaggaatg tctcccttgc cagtactgct agggtttttc
     7921 tttcaaacta tggaagccca ttcaagctgc atattacgat tttgtttttc gcttttagaa
     7981 agtggtttag atgagataat agaaaaattc ttgatctccg acaacgagta cttttatttt
     8041 ttttgctaat cactttactc aatattagct cgaaatcgta gaaacgtaga cgggtgcggg
     8101 ataccgagtg gtgtagttaa gaatttttat aaaccacgtg gcccaaaaat atgaacccaa
     8161 aacgtttata catgagtata ctttaagaag gctatacccc ttcgtgttag atgtagtttt
     8221 agctacccaa cccgagtcta tgagcttgac ttcagatgta gaaggcatta aatcgttttg
     8281 aatattaatt aaaaaacgat gaaaattaaa tatttaaaag caatcatacg ctgaaaattt
     8341 agtgctgtgg ctaatccttc aacatggaaa tgccataaaa gtgactttga caaaaaaaaa
     8401 agtatataca ggtagtaaac tcatctactt cattgacttt gtttacagca tgtggaagga
     8461 ggaatattta ttgctaaatc gtagtttaac attcaataag taatactatt gaaattcgac
     8521 aagattggcc gcatggatga aaaagaggca ttttgctttg ggagaattag ttcaaattag
     8581 aactgaaaaa aaaaacttta cgaggcaaaa atgtcggatt gagatcgtaa aagttcgctc
     8641 gtcgtctttt gctttgtgat tgttttcatg gatacatctt gctggatatt taaattttag
     8701 tactatgtat aagatattct ataaatgttt tatcacccaa acctgttagc gccttcttaa
     8761 ttctattcaa tctggctttt gctctgagac tacttcttgg actttcacta cttgttagtt
     8821 atacggaatt tgtgtaatta gaagtgaaat aatcctttct attagtaatg cgagctccaa
     8881 ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga
     8941 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     9001 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     9061 tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     9121 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     9181 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     9241 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     9301 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     9361 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     9421 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     9481 acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttcct gatgcggtat
     9541 tttctcctta cgcatctgtg cggtatttca caccgcatat ggtgcactct cagtacaatc
     9601 tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc
     9661 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc
     9721 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cga
//
