LOCUS       Exported                8008 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pVTLU263, complete sequence.
ACCESSION   AY974038
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8008)
  AUTHORS   Mok CSL., Melcher K.
  TITLE     Leu2d-based yeast expression vectors
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8008)
  AUTHORS   Mok CSL., Melcher K.
  TITLE     Direct Submission
  JOURNAL   Submitted (25-MAR-2005) School of Biomedical Sciences, University of
            Ulster, Cromore Road, Coleraine BT52 1SA, UK
REFERENCE   3  (bases 1 to 8008)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8008
                     /organism="Expression vector pVTLU263"
                     /lab_host="Saccharomyces cerevisiae"
                     /mol_type="other DNA"
                     /db_xref="taxon:322727"
     CDS             209..1303
                     /codon_start=1
                     /gene="LEU2"
                     /allele="leu2d"
                     /product="beta-isopropylmalate dehydrogenase"
                     /label=LEU2
                     /note="Leu2p"
                     /protein_id="AAX85435.1"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEALRYPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPGFAKNKVDPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     gene            209..1303
                     /gene="LEU2"
                     /allele="leu2d"
                     /label=LEU2
     primer_bind     1971..1987
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2038..2253
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             2254..3057
                     /codon_start=1
                     /gene="URA3-deltaNcoI"
                     /product="orotidine-5-phosphate decarboxylase"
                     /label=URA3-deltaNcoI
                     /note="Ura3p; NcoI site removed by site-specific 
                     mutagenesis without altering the amino acid sequence"
                     /protein_id="AAX85436.1"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     gene            2254..3057
                     /gene="URA3-deltaNcoI"
                     /label=URA3-deltaNcoI
     promoter        3255..3359
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3360..4220
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDVGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     CDS             3909..4220
                     /codon_start=1
                     /gene="ampR"
                     /product="AmpR"
                     /label=ampR
                     /note="beta-lactamase; antibiotic resistance marker"
                     /protein_id="AAX85437.1"
                     /translation="MATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWF
                     IADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW"
     gene            3909..4220
                     /gene="ampR"
                     /label=ampR
     rep_origin      4391..4979
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     rep_origin      4703
                     /label=Escherichia coli replication initiation base
                     /note="Escherichia coli replication initiation base"
     regulatory      5008..5409
                     /regulatory_class="promoter"
                     /note="ADH1"
     promoter        5008..5409
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             5418..5438
                     /codon_start=1
                     /product="His6-tag"
                     /label=His6-tag
                     /note="affinity purification tag"
                     /protein_id="AAX85438.1"
                     /translation="MHHHHHH"
     CDS             5421..5438
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             5448..6092
                     /codon_start=1
                     /product="glutathione-S-transferase"
                     /label=glutathione-S-transferase
                     /note="GST; affinity purification tag"
                     /protein_id="AAX85439.1"
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDH"
     misc_feature    6096..6119
                     /note="cleavage site for Tobacco Etch Virus (TEV) protease;
                     allows site-specific proteolytic cleavage of His6-GST 
                     tandem affinity tags"
     CDS             6096..6116
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     regulatory      6299..6491
                     /regulatory_class="terminator"
                     /note="ADH1 terminator"
     terminator      6299..6486
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     rep_origin      6497..7843
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     rep_origin      6543..7843
                     /label=from yeast endogeneous 2 micron plasmid
                     /note="from yeast endogeneous 2 micron plasmid"