LOCUS       pPIC3.        7764 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Pichia pastoris HIS4 vector for methanol-inducible intracellular 
            expression of a protein.
ACCESSION   .
VERSION     .
KEYWORDS    pPIC3
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7764)
  AUTHORS   Invitrogen (Life Technologies)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 7764)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     A start codon is present in the MCS upstream of the SnaBI site.
FEATURES             Location/Qualifiers
     source          1..7764
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        2..937
                     /label=AOX1 promoter
                     /note="inducible promoter, regulated by methanol"
     misc_feature    938..982
                     /label=MCS
                     /note="MCS"
                     /note="multiple cloning site"
     CDS             949..951
                     /codon_start=1
                     /product="start codon that should be utilized or removed
                     when inserting a gene"
                     /label=start codon that should be utilized or removed
                     /note="ATG"
                     /translation="M"
     terminator      1062..1308
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     CDS             complement(1721..4255)
                     /codon_start=1
                     /gene="Pichia pastoris HIS4"
                     /product="multifunctional enzyme, required for histidine 
                     biosynthesis"
                     /label=Pichia pastoris HIS4
                     /note="PpHIS4"
                     /note="auxotrophic marker for Pichia pastoris"
                     /translation="MTFPLLPAYASVAEFDNSLSLVGKAVFPYAADQLHNLIKFTQSTE
                     LQVNVQVESSVTEDQFEELIDNLLKLYNNGINEVILDLDLAERVVQRMIPGARVIYRTL
                     VDKVASLPANASIAVPFSSPLGDLKSFTNGGSRTVYAFSETAKLVDVTSTVASGIIPII
                     DARQLTTEYELSEDVKKFPVSEILLASLTTDRPDGLFTTLVADSSNYSLGLVYSSKKSI
                     PEAIRTQTGVYQSRRHGLWYKGATSGATQKLLGIELDCDGDCLKFVVEQTGVGFCHLER
                     TSCFGQSKGLRAMEATLWDRKSNAPEGSYTKRLFDDEVLLNAKIREEADELAEAKSKED
                     IAWECADLFYFALVRCAKYGVTLDEVERNLDMKSLKVTRRKGDAKPGYTKEQPKEESKP
                     KEVPSEGRIELCKIDVSKASSQEIEDALRRPIQKTEQIMELVKPIVDNVRQNGDKALLE
                     LTAKFDGVALKTPVLEAPFPEELMQLPDNVKRAIDLSIDNVRKFHEAQLTETLQVETCP
                     GVVCSRFARPIEKVGLYIPGGTAILPSTSLMLGVPAKVAGCKEIVFASPPKKDGTLTPE
                     VIYVAHKVGAKCIVLAGGAQAVAAMAYGTETVPKCDKIFGPGNQFVTAAKMMVQNDTSA
                     LCSIDMPAGPSEVLVIADKYADPDFVASDLLSQAEHGIDSQVILLAVDMTDKELARIED
                     AVHNQAVQLPRVEIVRKCIAHSTTLSVATYEQALEMSNQYAPEHLILQIENASSYVDQV
                     QHAGSVFVGAYSPESCGDYSSGTNHTLPTYGYARQYSGVNTATFQKFITSQDVTPEGLK
                     HIGQAVMDLAAVEGLDAHRNAVKVRMEKLGLI"
     misc_feature    4610..5366
                     /label=AOX1 3' fragment
                     /note="region downstream of Pichia pastoris AOX1 gene"
     misc_feature    5509..5649
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(5835..6423)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6597..7454)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(7455..7559)
                     /label=AmpR promoter
ORIGIN
        1 agatctaaca tccaaagacg aaaggttgaa tgaaaccttt ttgccatccg acatccacag
       61 gtccattctc acacataagt gccaaacgca acaggagggg atacactagc agcagaccgt
      121 tgcaaacgca ggacctccac tcctcttctc ctcaacaccc acttttgcca tcgaaaaacc
      181 agcccagtta ttgggcttga ttggagctcg ctcattccaa ttccttctat taggctacta
      241 acaccatgac tttattagcc tgtctatcct ggcccccctg gcgaggttca tgtttgttta
      301 tttccgaatg caacaagctc cgcattacac ccgaacatca ctccagatga gggctttctg
      361 agtgtggggt caaatagttt catgttcccc aaatggccca aaactgacag tttaaacgct
      421 gtcttggaac ctaatatgac aaaagcgtga tctcatccaa gatgaactaa gtttggttcg
      481 ttgaaatgct aacggccagt tggtcaaaaa gaaacttcca aaagtcgcca taccgtttgt
      541 cttgtttggt attgattgac gaatgctcaa aaataatctc attaatgctt agcgcagtct
      601 ctctatcgct tctgaacccc ggtgcacctg tgccgaaacg caaatgggga aacacccgct
      661 ttttggatga ttatgcattg tctccacatt gtatgcttcc aagattctgg tgggaatact
      721 gctgatagcc taacgttcat gatcaaaatt taactgttct aacccctact tgacagcaat
      781 atataaacag aaggaagctg ccctgtctta aacctttttt tttatcatca ttattagctt
      841 actttcataa ttgcgactgg ttccaattga caagcttttg attttaacga cttttaacga
      901 caacttgaga agatcaaaaa acaactaatt attcgaagga tccaaaccat ggaaaatacg
      961 tagaattccc tagggcggcc gcgaattaat tcgccttaga catgactgtt cctcagttca
     1021 agttgggcac ttacgagaag accggtcttg ctagattcta atcaagagga tgtcagaatg
     1081 ccatttgcct gagagatgca ggcttcattt ttgatacttt tttatttgta acctatatag
     1141 tataggattt tttttgtcat tttgtttctt ctcgtacgag cttgctcctg atcagcctat
     1201 ctcgcagctg atgaatatct tgtggtaggg gtttgggaaa atcattcgag tttgatgttt
     1261 ttcttggtat ttcccactcc tcttcagagt acagaagatt aagtgagaag ttcgtttgtg
     1321 caagcttatc gataagcttt aatgcggtag tttatcacag ttaaattgct aacgcagtca
     1381 ggcaccgtgt atgaaatcta acaatgcgct catcgtcatc ctcggcaccg tcaccctgga
     1441 tgctgtaggc ataggcttgg ttatgccggt actgccgggc ctcttgcggg atatcgtcca
     1501 ttccgacagc atcgccagtc actatggcgt gctgctagcg ctatatgcgt tgatgcaatt
     1561 tctatgcgca cccgttctcg gagcactgtc cgaccgcttt ggccgccgcc cagtcctgct
     1621 cgcttcgcta cttggagcca ctatcgacta cgcgatcatg gcgaccacac ccgtcctgtg
     1681 gatctatcga atctaaatgt aagttaaaat ctctaaataa ttaaataagt cccagtttct
     1741 ccatacgaac cttaacagca ttgcggtgag catctagacc ttcaacagca gccagatcca
     1801 tcactgcttg gccaatatgt ttcagtccct caggagttac gtcttgtgaa gtgatgaact
     1861 tctggaaggt tgcagtgtta actccgctgt attgacgggc atatccgtac gttggcaaag
     1921 tgtggttggt accggaggag taatctccac aactctctgg agagtaggca ccaacaaaca
     1981 cagatccagc gtgttgtact tgatcaacat aagaagaagc attctcgatt tgcaggatca
     2041 agtgttcagg agcgtactga ttggacattt ccaaagcctg ctcgtaggtt gcaaccgata
     2101 gggttgtaga gtgtgcaata cacttgcgta caatttcaac ccttggcaac tgcacagctt
     2161 ggttgtgaac agcatcttca attctggcaa gctccttgtc tgtcatatcg acagccaaca
     2221 gaatcacctg ggaatcaata ccatgttcag cttgagacag aaggtctgag gcaacgaaat
     2281 ctggatcagc gtatttatca gcaataacta gaacttcaga aggcccagca ggcatgtcaa
     2341 tactacacag ggctgatgtg tcattttgaa ccatcatctt ggcagcagta acgaactggt
     2401 ttcctggacc aaatattttg tcacacttag gaacagtttc tgttccgtaa gccatagcag
     2461 ctactgcctg ggcgcctcct gctagcacga tacacttagc accaaccttg tgggcaacgt
     2521 agatgacttc tggggtaagg gtaccatcct tcttaggtgg agatgcaaaa acaatttctt
     2581 tgcaaccagc aactttggca ggaacaccca gcatcaggga agtggaaggc agaattgcgg
     2641 ttccaccagg aatatagagg ccaactttct caataggtct tgcaaaacga gagcagacta
     2701 caccagggca agtctcaact tgcaacgtct ccgttagttg agcttcatgg aatttcctga
     2761 cgttatctat agagagatca atggctctct taacgttatc tggcaattgc ataagttcct
     2821 ctgggaaagg agcttctaac acaggtgtct tcaaagcgac tccatcaaac ttggcagtta
     2881 gttctaaaag ggctttgtca ccattttgac gaacattgtc gacaattggt ttgactaatt
     2941 ccataatctg ttccgttttc tggataggac gacgaagggc atcttcaatt tcttgtgagg
     3001 aggccttaga aacgtcaatt ttgcacaatt caatacgacc ttcagaaggg acttctttag
     3061 gtttggattc ttctttaggt tgttccttgg tgtatcctgg cttggcatct cctttccttc
     3121 tagtgacctt tagggacttc atatccaggt ttctctccac ctcgtccaac gtcacaccgt
     3181 acttggcaca tctaactaat gcaaaataaa ataagtcagc acattcccag gctatatctt
     3241 ccttggattt agcttctgca agttcatcag cttcctccct aattttagcg ttcaacaaaa
     3301 cttcgtcgtc aaataaccgt ttggtataag aaccttctgg agcattgctc ttacgatccc
     3361 acaaggtggc ttccatggct ctaagaccct ttgattggcc aaaacaggaa gtgcgttcca
     3421 agtgacagaa accaacacct gtttgttcaa ccacaaattt caagcagtct ccatcacaat
     3481 ccaattcgat acccagcaac ttttgagttg ctccagatgt agcaccttta taccacaaac
     3541 cgtgacgacg agattggtag actccagttt gtgtccttat agcctccgga atagactttt
     3601 tggacgagta caccaggccc aacgagtaat tagaagagtc agccaccaaa gtagtgaata
     3661 gaccatcggg gcggtcagta gtcaaagacg ccaacaaaat ttcactgaca gggaactttt
     3721 tgacatcttc agaaagttcg tattcagtag tcaattgccg agcatcaata atggggatta
     3781 taccagaagc aacagtggaa gtcacatcta ccaactttgc ggtctcagaa aaagcataaa
     3841 cagttctact accgccatta gtgaaacttt tcaaatcgcc cagtggagaa gaaaaaggca
     3901 cagcgatact agcattagcg ggcaaggatg caactttatc aaccagggtc ctatagataa
     3961 ccctagcgcc tgggatcatc ctttggacaa ctctttctgc caaatctagg tccaaaatca
     4021 cttcattgat accattattg tacaacttga gcaagttgtc gatcagctcc tcaaattggt
     4081 cctctgtaac ggatgactca acttgcacat taacttgaag ctcagtcgat tgagtgaact
     4141 tgatcaggtt gtgcagctgg tcagcagcat agggaaacac ggcttttcct accaaactca
     4201 aggaattatc aaactctgca acacttgcgt atgcaggtag caagggaaat gtcatacttg
     4261 aagtcggaca gtgagtgtag tcttgagaaa ttctgaagcc gtatttttat tatcagtgag
     4321 tcagtcatca ggagatcctc tacgccggac gcatcgtggc cggcatcacc ggcgccacag
     4381 gtgcggttgc tggcgcctat atcgccgaca tcaccgatgg ggaagatcgg gctcgccact
     4441 tcgggctcat gagcgcttgt ttcggcgtgg gtatggtggc aggccccgtg gccgggggac
     4501 tgttgggcgc catctccttg catgcaccat tccttgcggc ggcggtgctc aacggcctca
     4561 acctactact gggctgcttc ctaatgcagg agtcgcataa gggagagcgt cgagtatcta
     4621 tgattggaag tatgggaatg gtgatacccg cattcttcag tgtcttgagg tctcctatca
     4681 gattatgccc aactaaagca accggaggag gagatttcat ggtaaatttc tctgactttt
     4741 ggtcatcagt agactcgaac tgtgagacta tctcggttat gacagcagaa atgtccttct
     4801 tggagacagt aaatgaagtc ccaccaataa agaaatcctt gttatcagga acaaacttct
     4861 tgtttcgaac tttttcggtg ccttgaacta taaaatgtag agtggatatg tcgggtagga
     4921 atggagcggg caaatgctta ccttctggac cttcaagagg tatgtagggt ttgtagatac
     4981 tgatgccaac ttcagtgaca acgttgctat ttcgttcaaa ccattccgaa tccagagaaa
     5041 tcaaagttgt ttgtctacta ttgatccaag ccagtgcggt cttgaaactg acaatagtgt
     5101 gctcgtgttt tgaggtcatc tttgtatgaa taaatctagt ctttgatcta aataatcttg
     5161 acgagccaag gcgataaata cccaaatcta aaactctttt aaaacgttaa aaggacaagt
     5221 atgtctgcct gtattaaacc ccaaatcagc tcgtagtctg atcctcatca acttgagggg
     5281 cactatcttg ttttagagaa atttgcggag atgcgatatc gagaaaaagg tacgctgatt
     5341 ttaaacgtga aatttatctc aagatctctg cctcgcgcgt ttcggtgatg acggtgaaaa
     5401 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     5461 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac
     5521 ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt
     5581 gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     5641 cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     5701 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     5761 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     5821 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     5881 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     5941 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     6001 ctcccttcgg gaagcgtggc gctttctcaa tgctcacgct gtaggtatct cagttcggtg
     6061 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     6121 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     6181 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     6241 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     6301 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     6361 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     6421 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     6481 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     6541 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     6601 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     6661 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     6721 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     6781 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     6841 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     6901 gccattgctg caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     6961 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     7021 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     7081 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     7141 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     7201 ccggcgtcaa cacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     7261 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     7321 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     7381 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     7441 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     7501 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     7561 acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc
     7621 tataaaaata ggcgtatcac gaggcccttt cgtcttcaag aattaattct catgtttgac
     7681 agcttatcat cgataagctg actcatgttg gtattgtgaa atagacgcag atcgggaaca
     7741 ctgaaaaata acagttatta ttcg
//
