LOCUS       Exported                6574 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning and expression vector pCEV-G4-Km, complete sequence.
ACCESSION   KF366483
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6574)
  AUTHORS   Vickers CE, Bydder SF, Zhou Y, Nielsen LK.
  TITLE     Dual gene expression cassette vectors with antibiotic selection 
            markers for engineering in Saccharomyces cerevisiae
  JOURNAL   Microb. Cell Fact. 12 (1), 96 (2013)
  PUBMED    24161108
REFERENCE   2  (bases 1 to 6574)
  AUTHORS   Bydder SF, Vickers CE.
  TITLE     Direct Submission
  JOURNAL   Submitted (08-JUL-2013) AIBN, The University of Queensland, Crn 
            College and Cooper Roads (Bldg 75), St Lucia, QLD 4072, Australia
REFERENCE   3  (bases 1 to 6574)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6574
                     /organism="Cloning and expression vector pCEV-G4-Km"
                     /lab_host="Saccharomyces cerevisiae"
                     /mol_type="other DNA"
                     /db_xref="taxon:1412978"
     terminator      2..167
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     regulatory      complement(2..166)
                     /regulatory_class="terminator"
                     /note="ADH1 terminator from Saccharomyces cerevisiae"
     misc_feature    315..341
                     /label=FLAG tag
                     /note="FLAG tag"
     CDS             complement(315..338)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      complement(352..937)
                     /regulatory_class="promoter"
                     /note="HXT7 promoter from Saccharomyces cerevisiae"
     regulatory      947..1365
                     /regulatory_class="promoter"
                     /note="TEF1 promoter from Saccharomyces cerevisiae CEN.PK"
     promoter        947..1353
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        1373..1391
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    1406..1441
                     /label=C-myc tag
                     /note="C-myc tag"
     CDS             1409..1438
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     regulatory      1468..1657
                     /regulatory_class="terminator"
                     /note="CYC1 terminator from Saccharomyces cerevisiae"
     terminator      1468..1657
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     rep_origin      1844..2511
                     /label=pUC origin of replication for Escherichia coli
                     /note="pUC origin of replication for Escherichia coli"
     rep_origin      complement(1900..2488)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(2659..3519)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=bla
                     /note="confers ampicillin resistance in Escherichia coli"
                     /protein_id="AGZ03676.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(2659..3519)
                     /gene="bla"
                     /label=bla
     promoter        complement(3520..3624)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3651..4993
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     rep_origin      3653..4799
                     /label=2 micron yeast origin of replication
                     /note="2 micron yeast origin of replication"
     protein_bind    complement(4614..4661)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     misc_recomb     5013..5046
                     /label=loxP recognition sequence for Cre recombinase
                     /note="loxP recognition sequence for Cre recombinase"
     protein_bind    complement(5013..5046)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     regulatory      5047..5444
                     /regulatory_class="promoter"
                     /note="TEF promoter from Ashbya gossypii"
     gene            5101..6457
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        5101..5444
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             5445..6254
                     /codon_start=1
                     /gene="aphA1"
                     /product="aminoglycoside 3-phosphotransferase"
                     /label=aphA1
                     /note="confers kanamycin/neomycin resistance in Escherichia
                     coli, G418 resistance in Saccharomyces cerevisiae"
                     /protein_id="AGZ03677.1"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     gene            5445..6254
                     /gene="aphA1"
                     /label=aphA1
     regulatory      6255..6519
                     /regulatory_class="terminator"
                     /note="TEF terminator from Ashbya gossypii"
     terminator      6260..6457
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     misc_recomb     6520..6553
                     /label=loxP recognition sequence for Cre recombinase
                     /note="loxP recognition sequence for Cre recombinase"
     protein_bind    complement(6520..6553)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."