LOCUS       Exported               10878 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Yeast expression vector pPIC9-TSE, complete sequence.
ACCESSION   LT727317
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10878)
  AUTHORS   De Schamphelaire W, Olbrechts A, Meert J, Verhelst K, Roggeman 
            Fonseca M, Vanhoucke M, Beyaert R.
  TITLE     BCCM/LMBP Plasmid collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 10878)
  AUTHORS   De Schamphelaire W.
  TITLE     Direct Submission
  JOURNAL   Submitted (02-FEB-2017) BCCM/LMBP, Universiteit Gent, 
            Technologiepark 927, 9052, BELGIUM
REFERENCE   3  (bases 1 to 10878)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..10878
                     /organism="Yeast expression vector pPIC9-TSE"
                     /mol_type="other DNA"
                     /note="BCCM/LMBP Plasmid collection (Ghent 
                     University,Belgium) accession number LMBP 4166."
                     /db_xref="taxon:1945490"
     promoter        2..937
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 promoter
                     /note="inducible promoter, regulated by methanol"
     regulatory      7..772
                     /regulatory_class="promoter"
                     /note="AOX1 promoter"
     misc_feature    773..937
                     /label=5 UTR of AOX1
                     /note="5 UTR of AOX1"
     CDS             949..1215
                     /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             949..1191
                     /codon_start=1
                     /note="unnamed protein product; ppMF"
                     /protein_id="SJL88088.1"
                     /translation="MRFPSIFTAVLFAASSALAAPVNTTTEDETAQIPAEAVIGYSDLE
                     GDFDVAVLPFSNSTNNGLLFINTTIASIAAKEEGVS"
     CDS             1219..3141
                     /codon_start=1
                     /note="unnamed protein product; mature TS"
                     /protein_id="SJL88089.1"
                     /translation="MLAPGSSRVELFKRKNSTVPFEDKAGKVTERVVHSFRLPALVNVD
                     GVMVAIADARYDTSNDNSLIDTVAKYSVDDGETWETQIAIKNSRVSSVSRVVDPTVIVK
                     GNKLYVLVGSYYSSRSYWSSHGDARDWDILLAVGEVTKSIAGGKITASIKWGSPVSLKK
                     FFPAEMEGMHTNQFLGGAGVAIVASNGNLVYPVQVTNKRKQVFSKIFYSEDDGKTWKFG
                     KGRSDFGCSEPVALEWEGKLIINTRVDWKRRLVYESSDMGNTWVEAVGTLSRVWGPSPK
                     SDHPGSQSSFTAVTIEGMRVMLFTHPLNFKGRWLRDRLNLWLTDNQRIYNVGQVSIGDE
                     NSAYSSVLYKDDKLYCLHEINTDEVYSLVFARLVGELRIIKSVLRSWKNWDSHLSSICT
                     PADPAASSSESGCGPAVTTVGLVGFLSGNASQNVWEDAYRCVNASTANAERVRNGLKFA
                     GVGGGALWPVSQQGQNQRYRFANHAFTLVASVTIHEAPRAASPLLGASLDSSGGKKLLG
                     LSYDEKHQWQPIYGSTPVTPTGSWETGKRYHVVLTVANKIGSVYIDGELLEGSGQTVVP
                     DGRTPDISHFYVGGYGRSDMPTISHVTVNNVLLYNRQLNTEEIRTLFLSQDLIGTEAHM
                     DSSSDT"
     misc_feature    3151..3189
                     /label=E-tag
                     /note="E-tag"
     CDS             3199..4090
                     /codon_start=1
                     /note="unnamed protein product; fd g3 fragment"
                     /protein_id="SJL88090.1"
                     /translation="TVESCLAKPHTENSFTNVWKDDKTLDRYANYEGCLWNATGVVVCT
                     GDETQCYGTWVPIGLAIPENEGGGSEGGGSEGGGSEGGGTKPPEYGDTPIPGYTYINPL
                     DGTYPPGTEQNPANPNPSLEESQPLNTFMFQNNRFRNRQGALTVYTGTVTQGTDPVKTY
                     YQYTPVSSKAMYDAYWNGKFRDCAFHSGFNEDPFVCEYQGQSSDLPQPPVNAGGGSGGG
                     SGGGSEGGGSEGGGSEGGGSEGGGSGGGSGSGDFDYEKMANANKGAMTENADENALQSD
                     AKGKLDSVATDYGAAI"
     regulatory      4176..4435
                     /regulatory_class="terminator"
                     /note="AOX1 terminator"
     terminator      4176..4422
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     CDS             complement(4835..7369)
                     /codon_start=1
                     /gene="Pichia pastoris HIS4"
                     /product="multifunctional enzyme, required for histidine 
                     biosynthesis"
                     /label=PpHIS4
                     /note="auxotrophic marker for Pichia pastoris"
                     /translation="MTFPLLPAYASVAEFDNSLSLVGKAVFPYAADQLHNLIKFTQSTE
                     LQVNVQVESSVTEDQFEELIDNLLKLYNNGINEVILDLDLAERVVQRMIPGARVIYRTL
                     VDKVASLPANASIAVPFSSPLGDLKSFTNGGSRTVYAFSETAKLVDVTSTVASGIIPII
                     DARQLTTEYELSEDVKKFPVSEILLASLTTDRPDGLFTTLVADSSNYSLGLVYSSKKSI
                     PEAIRTQTGVYQSRRHGLWYKGATSGATQKLLGIELDCDGDCLKFVVEQTGVGFCHLER
                     TSCFGQSKGLRAMEATLWDRKSNAPEGSYTKRLFDDEVLLNAKIREEADELAEAKSKED
                     IAWECADLFYFALVRCAKYGVTLDEVERNLDMKSLKVTRRKGDAKPGYTKEQPKEESKP
                     KEVPSEGRIELCKIDVSKASSQEIEDALRRPIQKTEQIMELVKPIVDNVRQNGDKALLE
                     LTAKFDGVALKTPVLEAPFPEELMQLPDNVKRAIDLSIDNVRKFHEAQLTETLQVETCP
                     GVVCSRFARPIEKVGLYIPGGTAILPSTSLMLGVPAKVAGCKEIVFASPPKKDGTLTPE
                     VIYVAHKVGAKCIVLAGGAQAVAAMAYGTETVPKCDKIFGPGNQFVTAAKMMVQNDTSA
                     LCSIDMPAGPSEVLVIADKYADPDFVASDLLSQAEHGIDSQVILLAVDMTDKELARIED
                     AVHNQAVQLPRVEIVRKCIAHSTTLSVATYEQALEMSNQYAPEHLILQIENASSYVDQV
                     QHAGSVFVGAYSPESCGDYSSGTNHTLPTYGYARQYSGVNTATFQKFITSQDVTPEGLK
                     HIGQAVMDLAAVEGLDAHRNAVKVRMEKLGLI"
     CDS             complement(4838..7369)
                     /codon_start=1
                     /note="unnamed protein product; Pichia HIS4"
                     /protein_id="SJL88091.1"
                     /translation="MTFPLLPAYASVAEFDNSLSLVGKAVFPYAADQLHNLIKFTQSTE
                     LQVNVQVESSVTEDQFEELIDNLLKLYNNGINEVILDLDLAERVVQRMIPGARVIYRTL
                     VDKVASLPANASIAVPFSSPLGDLKSFTNGGSRTVYAFSETAKLVDVTSTVASGIIPII
                     DARQLTTEYELSEDVKKFPVSEILLASLTTDRPDGLFTTLVADSSNYSLGLVYSSKKSI
                     PEAIRTQTGVYQSRRHGLWYKGATSGATQKLLGIELDCDGDCLKFVVEQTGVGFCHLER
                     TSCFGQSKGLRAMEATLWDRKSNAPEGSYTKRLFDDEVLLNAKIREEADELAEAKSKED
                     IAWECADLFYFALVRCAKYGVTLDEVERNLDMKSLKVTRRKGDAKPGYTKEQPKEESKP
                     KEVPSEGRIELCKIDVSKASSQEIEDALRRPIQKTEQIMELVKPIVDNVRQNGDKALLE
                     LTAKFDGVALKTPVLEAPFPEELMQLPDNVKRAIDLSIDNVRKFHEAQLTETLQVETCP
                     GVVCSRFARPIEKVGLYIPGGTAILPSTSLMLGVPAKVAGCKEIVFASPPKKDGTLTPE
                     VIYVAHKVGAKCIVLAGGAQAVAAMAYGTETVPKCDKIFGPGNQFVTAAKMMVQNDTSA
                     LCSIDMPAGPSEVLVIADKYADPDFVASDLLSQAEHGIDSQVILLAVDMTDKELARIED
                     AVHNQAVQLPRVEIVRKCIAHSTTLSVATYEQALEMSNQYAPEHLILQIENASSYVDQV
                     QHAGSVFVGAYSPESCGDYSSGTNHTLPTYGYARQYSGVNTATFQKFITSQDVTPEGLK
                     HIGQAVMDLAAVEGLDAHRNAVKVRMEKLGLI"
     misc_feature    7724..8481
                     /label=3 flanking region of AOX1
                     /note="3 flanking region of AOX1"
     misc_feature    7724..8480
                     /label=AOX1 3 fragment
                     /note="region downstream of Pichia pastoris AOX1 gene"
     misc_feature    8623..8763
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(8765..9167)
                     /direction=LEFT
                     /label=pMB1 ori
                     /note="pMB1 ori"
     rep_origin      complement(8949..9537)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(9708..10568)
                     /codon_start=1
                     /note="unnamed protein product; ampicillin resistance"
                     /protein_id="SJL88092.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(10569..10673)
                     /gene="bla"
                     /label=AmpR promoter