LOCUS       40924_35414        7238 bp DNA     circular SYN 18-DEC-2018
DEFINITION  E.coli-K.pastoris shuttle vector pPpHIS4, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7238)
  AUTHORS   Naatsaari L, Mistlberger B, Ruth C, Hajek T, Hartner FS, Glieder A.
  TITLE     Deletion of the Pichia pastoris KU70 Homologue Facilitates Platform 
            Strain Generation for Gene Expression and Synthetic Biology
  JOURNAL   PLoS ONE 7 (6), E39720 (2012)
  PUBMED    22768112
REFERENCE   2  (bases 1 to 7238)
  AUTHORS   Naeaetsaari L, Mistlberger B, Ruth C, Hajek T, Hartner F, Glieder A.
  TITLE     Direct Submission
  JOURNAL   Submitted (30-JAN-2012) Institute of Molecular Biotechnology, Graz 
            University of Technology, Petersgasse 14/2, Graz, Styria 8010, 
            Austria
REFERENCE   3  (bases 1 to 7238)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7238)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PLoS ONE"; 
            date: "2012"; volume: "7"; issue: "6"; pages: "E39720"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (30-JAN-2012) Institute of Molecular Biotechnology, Graz University 
            of Technology, Petersgasse 14/2, Graz, Styria 8010, Austria"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Assembly Method       :: SeqMan v. 7.0.0
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..7238
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        134..1068
                     /label=AOX1 promoter
                     /note="inducible promoter, regulated by methanol"
     terminator      1097..1343
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        1506..2210
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             2223..4757
                     /codon_start=1
                     /product="HIS4"
                     /label=HIS4
                     /note="HIS4; codon optimized"
                     /protein_id="AFJ20662.1"
                     /translation="MTFPLLPAYASVAEFDNSLSLVGKAVFPYAADQLHNLIKFTQSTE
                     LQVNVQVESSVTEDQFEELIDNLLKLYNNGINEVILDLDLAERVVQRMIPGARVIYRTL
                     VDKVASLPANASIAVPFSSPLGDLKSFTNGGSRTVYAFSETAKLVDVTSTVASGIIPII
                     DARQLTTEYELSEDVKKFPVSEILLASLTTDRPDGLFTTLVADSSNYSLGLVYSSKKSI
                     PEAIRTQTGVYQSRRHGLWYKGATSGATQKLLGIELDCDGDCLKFVVEQTGVGFCHLER
                     TSCFGQSKGLRAMEATLWDRKSNAPEGSYTKRLFDDEVLLNAKIREEADELAEAKSKED
                     IAWECADLFYFALVRCAKYGVTLDEVERNLDMKSLKVTRRKGDAKPGYTKEQPKEESKP
                     KEVPSEGRIELCKIDVSKASSQEIEDALRRPIQKTEQIMELVKPIVDNVRQNGDKALLE
                     LTAKFDGVALKTPVLEAPFPEELMQLPDNVKRAIDLSIDNVRKFHEAQLTETLQVETCP
                     GVVCSRFARPIEKVGLYIPGGTAILPSTSLMLGVPAKVAGCKEIVFASPPKKDGTLTPE
                     VIYVAHKVGAKCIVLAGGAQAVAAMAYGTETVPKCDKIFGPGNQFVTAAKMMVQNDTSA
                     LCSIDMPAGPSEVLVIADKYADPDFVASDLLSQAEHGIDSQVILLAVDMTDKELARIED
                     AVHNQAVQLPRVEIVRKCIAHSTTLSVATYEQALEMSNQYAPEHLILQIENASSYVDQV
                     QHAGSVFVGAYSPESCGDYSSGTNHTLPTYGYARQYSGVNTATFQKFITSQDVTPEGLK
                     HIGQAVMDLAAVEGLDAHRNAVKVRMEKLGLI"
     regulatory      4758..5289
                     /label=TIF51A terminator
                     /note="TIF51A terminator"
                     /regulatory_class="terminator"
     regulatory      5290..5347
                     /label=EM72 synthetic promoter
                     /note="EM72 synthetic promoter"
                     /regulatory_class="promoter"
     CDS             5354..6211
                     /label=AmpR
                     /note="beta-lactamase"
     terminator      6221..6408
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae
                     alcohol dehydrogenase 1 (ADH1) gene"
     rep_origin      6572..7160
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 agatccaatt cccgctttga ctgcctgaaa tctccatcgc ctacaatgat gacatttgga
       61 tttggttgac tcatgttggt attgtgaaat agacgcagat cgggaacact gaaaaataca
      121 cagttattat tcagatctaa catccaaaga cgaaaggttg aatgaaacct ttttgccatc
      181 cgacatccac aggtccattc tcacacataa gtgccaaacg caacaggagg ggatacacta
      241 gcagcagacc gttgcaaacg caggacctcc actcctcttc tcctcaacac ccacttttgc
      301 catcgaaaaa ccagcccagt tattgggctt gattggagct cgctcattcc aattccttct
      361 attaggctac taacaccatg actttattag cctgtctatc ctggcccccc tggcgaggtt
      421 catgtttgtt tatttccgaa tgcaacaagc tccgcattac acccgaacat cactccagat
      481 gagggctttc tgagtgtggg gtcaaatagt ttcatgttcc ccaaatggcc caaaactgac
      541 agtttaaacg ctgtcttgga acctaatatg acaaaagcgt gatctcatcc aagatgaact
      601 aagtttggtt cgttgaaatg ctaacggcca gttggtcaaa aagaaacttc caaaagtcgg
      661 cataccgttt gtcttgtttg gtattgattg acgaatgctc aaaaataatc tcattaatgc
      721 ttagcgcagt ctctctatcg cttctgaacc ccggtgcacc tgtgccgaaa cgcaaatggg
      781 gaaacacccg ctttttggat gattatgcat tgtctccaca ttgtatgctt ccaagattct
      841 ggtgggaata ctgctgatag cctaacgttc atgatcaaaa tttaactgtt ctaaccccta
      901 cttgacagca atatataaac agaaggaagc tgccctgtct taaccttttt ttttatcatc
      961 attattagct tactttcata attgcgactg gttccaattg acaagctttt gattttaacg
     1021 acttttaacg acaacttgag aagatcaaaa aacaactaat tattgaaaga attcactagt
     1081 ggcgcgccgc ggccgctcaa gaggatgtca gaatgccatt tgcctgagag atgcaggctt
     1141 catttttgat acttttttat ttgtaaccta tatagtatag gatttttttt gtcattttgt
     1201 ttcttctcgt acgagcttgc tcctgatcag cctatctcgc agcagatgaa tatcttgtgg
     1261 taggggtttg ggaaaatcat tcgagtttga tgtttttctt ggtatttccc actcctcttc
     1321 agagtacaga agattaagtg agaccttcgt ttgtgcggat cctttattat gttacaatat
     1381 ggaagggaac tttacacttc tcctatgcac atatattaat taaagtccaa tgctagtaga
     1441 gaaggggggt aacacccctc cgcgctcttt tccgattttt ttctaaaccg tggaatattt
     1501 cggatatcct tttgttgttt ccgggtgtac aatatggact tcctcttttc tggcaaccaa
     1561 acccatacat cgggattcct ataatacctt cgttggtctc cctaacatgt aggtggcgga
     1621 ggggagatat acaatagaac agataccaga caagacataa tgggctaaac aagactacac
     1681 caattacact gcctcattga tggtggtaca taacgaacta atactgtagc cctagacttg
     1741 atagccatca tcatatcgaa gtttcactac cctttttcca tttgccatct attgaagtaa
     1801 taataggcgc atgcaacttc ttttcttttt ttttcttttc tctctccccc gttgttgtct
     1861 caccatatcc gcaatgacaa aaaaatgatg gaagacacta aaggaaaaaa ttaacgacaa
     1921 agacagcacc aacagatgtc gttgttccag agctgatgag gggtatctcg aagcacacga
     1981 aactttttcc ttccttcatt cacgcacact actctctaat gagcaacggt atacggcctt
     2041 ccttccagtt acttgaattt gaaataaaaa aaagtttgct gtcttgctat caagtataaa
     2101 tagacctgca attattaatc ttttgtttcc tcgtcattgt tctcgttccc tttcttcctt
     2161 gtttcttttt ctgcacaata tttcaagcta taccaagcat acaatcaact atctcatata
     2221 caatgacctt tcccttgctt ccagcctacg cttctgtcgc cgagtttgac aactccctta
     2281 gcttggttgg caaagccgtc tttccctatg cagccgatca acttcacaac ttgatcaagt
     2341 tcacacagtc caccgagctt caagtcaatg tgcaagttga atcgtctgtt acggaagacc
     2401 agtttgaaga gcttatcgac aaccttttga agctttacaa caatggtatc aacgaggtca
     2461 ttcttgacct tgatcttgct gaaagagttg tgcaacgaat gattcctgga gctagagtca
     2521 tctacagaac ccttgttgac aaagttgctt cacttcccgc taacgcctct attgctgttc
     2581 ctttctcttc accccttggt gatcttaagt cattcacaaa cggtggaagc cgtaccgtgt
     2641 atgctttctc ggagacggcc aaacttgttg acgttacctc tacagttgct tctggaatca
     2701 ttcctatcat tgatgcacga caacttacta cagagtatga actttccgag gatgtgaaga
     2761 agttccccgt ctctgagatc cttttggctt ctcttaccac agaccgtcca gatggtttgt
     2821 tcactaccct tgttgccgat tcttccaact actcacttgg tcttgtgtac tcttccaaga
     2881 aatccatccc cgaggctatc agaactcaga ccggtgtcta tcaatccaga cgacatggac
     2941 tttggtacaa aggtgccaca tccggtgcaa cacagaagtt gcttggcatt gagcttgact
     3001 gtgatggaga ttgtcttaag tttgtggttg agcaaactgg tgtcggattc tgtcatcttg
     3061 aacgaacgtc ttgctttgga caatccaaag gccttagagc catggaggct acactttggg
     3121 atcgtaagtc caatgcacca gagggttcgt acaccaaacg attgtttgac gatgaggtgt
     3181 tgcttaacgc caagattcga gaagaggctg atgaacttgc agaagctaag tccaaagagg
     3241 acattgcatg ggaatgtgcc gatttgttct actttgccct tgttagatgc gcaaagtatg
     3301 gagttactct tgatgaagtg gaacgtaacc ttgatatgaa gtcccttaag gttactcgac
     3361 gtaaaggtga tgcaaagcca ggctacacta aggaacaacc caaagaggaa tcgaagccca
     3421 aagaggtgcc ttccgaaggc agaatcgaac tttgcaagat tgatgtttct aaggcttcgt
     3481 ctcaagagat tgaagatgcc cttcgtcgac caattcagaa aactgaacag atcatggagc
     3541 ttgtgaaacc aatcgtcgac aatgttcgtc agaatggtga caaagccttg cttgagttga
     3601 ctgccaagtt cgatggagtc gccttgaaaa cccctgtgct tgaagctcca ttccctgagg
     3661 aacttatgca gcttcccgac aatgtgaaac gtgcaattga tctttccatt gacaatgtta
     3721 gaaagtttca tgaagcacaa cttactgaga cgcttcaagt tgaaacttgt ccaggagtcg
     3781 tgtgctcacg attcgctaga cctatcgaga aagtcggcct ttacattcct ggcggaacgg
     3841 ctatccttcc tagcacctct cttatgcttg gagttcctgc caaagtcgca ggatgtaagg
     3901 agattgtgtt tgcctctcca cctaagaaag atggcacact tactcctgag gtcatctatg
     3961 tcgctcacaa agttggtgcc aagtgcattg ttcttgctgg aggtgcacaa gctgttgctg
     4021 caatggccta tggtactgaa accgttccca agtgtgacaa gattttcggt cctggaaacc
     4081 agtttgttac agcagctaag atgatggttc agaatgatac ctcggccctt tgttccatcg
     4141 acatgcccgc tggaccaagc gaagttcttg tcattgccga caagtatgct gatcctgact
     4201 ttgtggcctc tgacttgctt tctcaagctg aacatggcat tgattcccag gtgatcttgc
     4261 ttgcagtgga catgaccgac aaggagcttg cacgaattga agatgccgtt cacaatcagg
     4321 cagtgcagct tcccagagtc gagattgttc gaaagtgcat tgcacattct actacactta
     4381 gcgttgctac ctacgaacag gctcttgaga tgtctaacca gtatgctcct gaacacttga
     4441 ttcttcagat tgagaatgcc tcttcctatg tcgatcaagt ccaacatgcc ggttccgtgt
     4501 ttgtcggcgc atactcacct gagtcctgtg gtgactacag ctctggaacc aatcatacgc
     4561 ttcctactta cggctatgct agacagtact ctggagtgaa cactgctacc tttcagaagt
     4621 tcatcacttc tcaagatgtt actccagaag gtttgaaaca cattggtcaa gccgtcatgg
     4681 accttgcagc tgttgaaggt cttgatgctc accgtaacgc tgtcaaagtt cgaatggaga
     4741 agcttggttt gatttaaggc cgcaccggtt aacatcatgg catgggatat aaatgaaaaa
     4801 agaaaaaaaa actccgacgc cccttccatc acatcatgta ctcttcgctg aaccgggttt
     4861 ttttctttgc aatttttttt tcgttctcct aaagcataca caaataaatc ctttttttta
     4921 ttttctattt attttgttat ttatcatcta tatagcaata atatactttg tttttattcg
     4981 tatttcacac ttttcttttt ccttatgcag gcagtgtaat tcattgggga ggatgatttt
     5041 catgtgcgca tatctaccgg ctgcaagcag ccggtcggtg gcaaatccgg cgcttccccc
     5101 tcaaaaaaaa aaaaaaaaaa aagggaactc tcagaacggg ggaggttgaa gagcaggcca
     5161 agggaaatat tagttttgac ctatgtggga aacagaattt tcaatgagtt atggcaactt
     5221 ggccgagtgg ttaaggcgaa agattagaaa tcttttgggc tttgcccgcg caggttcgag
     5281 tcctgcaggg tgttgacact ttatacttcc ggctcgtata atacgacaag gtgtaaggag
     5341 gactaaatct agaatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     5401 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     5461 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     5521 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     5581 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     5641 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     5701 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     5761 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     5821 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     5881 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     5941 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     6001 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     6061 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     6121 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     6181 tgagataggt gcctcactga ttaagcattg gtaacctagg gcgaatttct tatgatttat
     6241 gatttttatt attaaataag ttataaaaaa aataagtgta tacaaatttt aaagtgactc
     6301 ttaggtttta aaacgaaaat tcttattctt gagtaactct ttcctgtagg tcaggttgct
     6361 ttctcaggta tagcatgagg tcgctcttat tgaccacacc tctaccggca tgccgagcaa
     6421 atgcctgcaa atcgctcccc atttcaccca attgtagata tgctaactcc agcaatgagt
     6481 tgatgaatct cggtgtgtat tttatgtcct cagaggacaa ctgcaggtac cactgagcgt
     6541 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     6601 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     6661 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     6721 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     6781 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     6841 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     6901 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     6961 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     7021 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     7081 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     7141 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     7201 gctggccttt tgctcacatg ttctttcctg cggtaccc
//
