LOCUS       Exported                6834 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Mammalian expression vector pCBio, complete sequence.
ACCESSION   DQ520291
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6834)
  AUTHORS   Kulman JD, Satake M, Harris JE.
  TITLE     A versatile system for site-specific enzymatic biotinylation and 
            regulated expression of proteins in cultured mammalian cells
  JOURNAL   Protein Expr. Purif. 52 (2), 320-328 (2007)
  PUBMED    17084093
REFERENCE   2  (bases 1 to 6834)
  AUTHORS   Kulman JD, Satake M, Harris JE.
  TITLE     Direct Submission
  JOURNAL   Submitted (28-APR-2006) Biochemistry, University of Washington, N.E.
            Pacific St., Seattle, WA 98195, USA
REFERENCE   3  (bases 1 to 6834)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6834
                     /organism="Mammalian expression vector pCBio"
                     /mol_type="other DNA"
                     /db_xref="taxon:386595"
     regulatory      7..439
                     /regulatory_class="promoter"
                     /note="tetracycline-responsive promoter"
     protein_bind    15..285
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     protein_bind    15..33
                     /label=TetR binding site
                     /bound_moiety="TetR"
                     /note="tetO repeat"
     protein_bind    57..75
                     /label=TetR binding site
                     /bound_moiety="TetR"
                     /note="tetO repeat"
     protein_bind    99..117
                     /label=TetR binding site
                     /bound_moiety="TetR"
                     /note="tetO repeat"
     protein_bind    141..159
                     /label=TetR binding site
                     /bound_moiety="TetR"
                     /note="tetO repeat"
     protein_bind    183..201
                     /label=TetR binding site
                     /bound_moiety="TetR"
                     /note="tetO repeat"
     protein_bind    225..243
                     /label=TetR binding site
                     /bound_moiety="TetR"
                     /note="tetO repeat"
     protein_bind    267..285
                     /label=TetR binding site
                     /bound_moiety="TetR"
                     /note="tetO repeat"
     promoter        318..356
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     misc_feature    320..439
                     /label=minimal CMV promoter
                     /note="minimal CMV promoter"
     regulatory      342..349
                     /regulatory_class="TATA_box"
     misc_feature    471..490
                     /note="multiple cloning site; MCS"
     misc_feature    491..529
                     /label=biotin acceptor peptide (BioTag)-encoding
                     /note="biotin acceptor peptide (BioTag)-encoding"
     misc_feature    584..1164
                     /note="encephalomyocarditis virus internal ribosome entry 
                     site; IRES"
     misc_feature    591..1164
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     gene            1209..2222
                     /gene="birA"
                     /label=birA
     regulatory      1209..1217
                     /regulatory_class="ribosome_binding_site"
                     /gene="birA"
                     /note="Kozak site"
     regulatory      1212..1221
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1218..2222
                     /codon_start=1
                     /gene="birA"
                     /product="biotin ligase"
                     /label=birA
                     /note="holocarboxylase synthetase"
                     /protein_id="ABF74577.1"
                     /translation="MDYKDDDDKSPRSMKDNTVPLKLIALLANGEFHSGEQLGETLGMS
                     RAAINKHIQTLRDWGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTN
                     QYLLDRIGELKSGDACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLS
                     LVIGIVMAEVLRKLGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINM
                     AMRRVEESVVNQGWITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKL
                     DNFINRPVKLIIGDKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     misc_feature    1221..1244
                     /gene="birA"
                     /label=FLAG epitope tag-encoding
                     /note="FLAG epitope tag-encoding"
     CDS             1221..1244
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    1257..2222
                     /gene="birA"
                     /label=Escherichia coli BirA (biotin ligase)
                     /note="Escherichia coli BirA (biotin ligase)"
     CDS             1257..2222
                     /codon_start=1
                     /gene="birA"
                     /product="E. coli biotin protein ligase"
                     /label=BirA
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK
                     LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG
                     WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG
                     DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     regulatory      2272..3439
                     /regulatory_class="polyA_signal_sequence"
                     /note="beta-globin poly-A signal"
     intron          2287..2859
                     /label=-beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     polyA_site      3056..3111
                     /label=-beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal"
     rep_origin      complement(3673..4261)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4432..5292)
                     /codon_start=1
                     /gene="amp"
                     /product="beta-lactamase"
                     /label=amp
                     /note="confers ampicillin resistance"
                     /protein_id="ABF74578.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(4432..5292)
                     /gene="amp"
                     /label=amp
     promoter        complement(5293..5397)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5502..5831
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     regulatory      5515..5923
                     /regulatory_class="promoter"
                     /note="SV40 promoter"
     rep_origin      5682..5817
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5924..6523
                     /codon_start=1
                     /gene="pur"
                     /product="puromycin N-acetyltransferase"
                     /label=pur
                     /note="confers puromycin resistance"
                     /protein_id="ABF74579.1"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     gene            5924..6523
                     /gene="pur"
                     /label=pur
     regulatory      6524..6834
                     /regulatory_class="polyA_signal_sequence"
                     /note="SV40 poly-A signal"
     polyA_signal    6753..6834
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"