LOCUS       Exported                4694 bp ds-DNA    circular SYN 06-2-2013
DEFINITION  Pichia pastoris vector for methylamine-inducible expression of a 
            secreted protein.
ACCESSION   .
VERSION     .
KEYWORDS    pFLD(alpha)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4694)
  AUTHORS   Invitrogen (Life Technologies)
  TITLE     Direct Submission
COMMENT     Linearize within the FLD1 promoter for integration.
FEATURES             Location/Qualifiers
     source          1..4694
                     /organism="synthetic DNA construct"
                     /lab_host="Pichia pastoris"
                     /mol_type="other DNA"
     promoter        1..597
                     /gene="Pichia pastoris FLD1"
                     /note="FLD1 promoter"
                     /note="methanol- and methylamine-inducible promoter for 
                     Pichia pastoris formaldehyde dehydrogenase"
     CDS             622..885
                     /codon_start=1
                     /gene="MF-alpha-1"
                     /product="N-terminal secretion signal from S. cerevisiae 
                     alpha-factor"
                     /note="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
                     GDFDVAVLPFSNSTNNGLLFINTTIASIAAKEEGVSLEKREAE"
     misc_feature    889..959
                     /note="MCS"
                     /note="multiple cloning site"
     CDS             964..1005
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /note="V5 tag"
                     /translation="GKPIPNPLLGLDST"
     CDS             1015..1032
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /note="6xHis"
                     /translation="HHHHHH"
     terminator      1112..1358
                     /gene="Pichia pastoris AOX1"
                     /note="AOX1 terminator"
                     /note="transcription terminator for AOX1"
     promoter        1725..1829
                     /gene="bla"
                     /note="AmpR promoter"
     CDS             1830..2690
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      2861..3449
                     /direction=RIGHT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     promoter        3504..3910
                     /gene="S. cerevisiae TEF1"
                     /note="TEF1 promoter"
                     /note="promoter for EF-1-alpha"
     promoter        3917..3964
                     /note="EM7 promoter"
                     /note="synthetic bacterial promoter "
     CDS             3983..4357
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /note="BleoR"
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGQ
                     EFALRDPAGNCVHFVAEEQD"
     terminator      4423..4670
                     /gene="S. cerevisiae CYC1"
                     /note="CYC1 terminator"
                     /note="transcription terminator for CYC1"