LOCUS       Exported                3551 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pPICZalphaH6E, complete sequence.
ACCESSION   KM035419
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3551)
  AUTHORS   Janakiraman VN, Cabanne C, Dieryck W, Hocquellet A, Joucla G, Le 
            Senechal C, Chaignepain S, Costaglioli P, Santarelli X, Garbay B, 
            Noubhani A.
  TITLE     Production and purification of recombinant human hepcidin-25 with 
            authentic N and C-termini
  JOURNAL   J. Biotechnol. 195, 89-92 (2015)
  PUBMED    25562424
REFERENCE   2  (bases 1 to 3551)
  AUTHORS   Dieryck W, Narasimhan Janakiraman V, Cabanne C, Hocquellet A, Joucla
            G, Le Senechal C, Chaignepain S, Costaglioli P, Santarelli X, Garbay
            B, Noubhani A.
  TITLE     Direct Submission
  JOURNAL   Submitted (19-JUN-2014) ENSTBB, Institut Polytechnique de Bordeaux, 
            146 rue Leo Saignat, Bordeaux, gironde 33076, France
REFERENCE   3  (bases 1 to 3551)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..3551
                     /organism="Expression vector pPICZalphaH6E"
                     /host="Pichia pastoris"
                     /mol_type="other DNA"
                     /db_xref="taxon:1546260"
     misc_feature    1..941
                     /label=5 AOX1 promoter region
                     /note="5 AOX1 promoter region"
     promoter        2..940
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 promoter
                     /note="inducible promoter, regulated by methanol"
     CDS             941..1354
                     /codon_start=1
                     /product="alpha factor secretion signal/UV-Tag/6xTag fusion
                     protein"
                     /label=alpha factor secretion signal/UV-Tag/6xTag fusi
                     /protein_id="AIP93596.1"
                     /translation="MRFPSIFTAVLFAASSALAAPVNTTTEDETAQIPAEAVIGYSDLE
                     GDFDVAVLPFSNSTNNGLLFINTTIASIAAKEEGVSLEKREAEAELWSPSYYASHHHHH
                     HSSSNNNNNNNNNNLGDDDDKVPEFACSRVDLQ"
     misc_feature    941..1207
                     /label=alpha factor secretion sequence
                     /note="alpha factor secretion sequence"
     CDS             941..1207
                     /codon_start=1
                     /gene="MF-alpha-1"
                     /product="N-terminal secretion signal from S. cerevisiae 
                     alpha-factor"
                     /label=-alpha-factor secretion signal
                     /note="Cleavage by the Kex2 protease occurs after the 
                     dibasic KR sequence. The EA dipeptides are then removed by 
                     dipeptidyl aminopeptidase A."
                     /translation="MRFPSIFTAVLFAASSALAAPVNTTTEDETAQIPAEAVIGYSDLE
                     GDFDVAVLPFSNSTNNGLLFINTTIASIAAKEEGVSLEKREAEA"
     CDS             1238..1255
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    1301..1315
                     /label=enterokinase cleavage site
                     /note="enterokinase cleavage site"
     CDS             1301..1315
                     /codon_start=1
                     /product="enterokinase recognition and cleavage site"
                     /label=enterokinase site
                     /translation="DDDDK"
     misc_feature    1315..1359
                     /label=multiple cloning site
                     /note="multiple cloning site"
     regulatory      1360..1640
                     /regulatory_class="terminator"
                     /note="AOX1 transcription termination region"
     terminator      1375..1621
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        1636..2047
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     misc_feature    1641..2051
                     /label=TEF1 promoter
                     /note="TEF1 promoter"
     misc_feature    2053..2120
                     /label=EM7 promoter
                     /note="EM7 promoter"
     promoter        2055..2102
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2121..2495
                     /codon_start=1
                     /product="zeocin resistance protein"
                     /label=zeocin resistance protein
                     /note="Sh ble"
                     /protein_id="AIP93597.1"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     regulatory      2496..2813
                     /regulatory_class="terminator"
                     /note="CYC1 transciption terminator region"
     terminator      2561..2808
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     rep_origin      complement(2865..3493)
                     /direction=LEFT
                     /label=pUC origin of replication
                     /note="pUC origin of replication"
     rep_origin      complement(2883..3471)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"