LOCUS       Exported                9654 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  PGREG expression vector pGREG505-G, complete sequence.
ACCESSION   HM592665
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9654)
  AUTHORS   Karreman RJ, Morrissey JP.
  TITLE     Controlling expression of genes in Saccharomyces cerevisiae using 
            variants of the pGREG recombinational vectors
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 9654)
  AUTHORS   Karreman RJ, Morrissey JP.
  TITLE     Direct Submission
  JOURNAL   Submitted (28-JUN-2010) Microbiology Department, University College 
            Cork, College Rd, Cork City, Cork 0000, Ireland
REFERENCE   3  (bases 1 to 9654)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..9654
                     /organism="pGREG expression vector pGREG505-G"
                     /mol_type="other DNA"
                     /PCR_primers="fwd_name: Greg-Seq-F, fwd_seq:
                     gaattcgatatcaagcttatc, rev_name: Greg-Seq-R, rev_seq: 
                     gtgacataactaattacatg"
                     /db_xref="taxon:913764"
     rep_origin      7..462
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        762..1166
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             1167..2273
                     /codon_start=1
                     /product="isopropylmalate dehydrogenase"
                     /label=isopropylmalate dehydrogenase
                     /note="Saccharomyces cerevisiae Leu2p"
                     /protein_id="ADP69179.1"
                     /translation="MSAPMSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDF
                     ENHLIGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKEL
                     QLYANLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDS
                     EQYTVPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLK
                     VQHQLIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDK
                     NTAFGLYEPCHGSAPDLPKNKVDPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAG
                     IRTGDLGGSNSTTEVGDAVAEEVKKILA"
     CDS             1179..2273
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVDPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     rep_origin      3000..3518
                     /label=CEN/ARS plasmid origin
                     /note="CEN/ARS plasmid origin"
     misc_feature    3005..3508
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        3545..3649
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3650..4510
                     /codon_start=1
                     /product="beta-lactamase"
                     /label=beta-lactamase
                     /note="ampicillin resistance protein"
                     /protein_id="ADP69180.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      4681..5269
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     promoter        5593..5623
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5631..5647
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     5655..5671
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        5692..5710
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      5729..6408
                     /regulatory_class="promoter"
                     /note="Saccharomyces cerevisiae GPD1 promoter (intermediate
                     strength)"
     misc_feature    6433..6466
                     /note="Rec1 site; pGREG recombinational cloning site I"
     misc_feature    6460..6465
                     /label=SalI restriction enzyme cut site
                     /note="SalI restriction enzyme cut site"
     misc_feature    6468..7627
                     /label=HIS3 cassette (excised by SalI)
                     /note="HIS3 cassette (excised by SalI)"
     promoter        6755..6942
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             6943..7602
                     /codon_start=1
                     /product="imidazoleglycerol-phosphate dehydratase"
                     /label=imidazoleglycerol-phosphate dehydratase
                     /note="Saccharomyces cerevisiae His3p"
                     /protein_id="ADP69181.1"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALLARGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHFL
                     ESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     misc_feature    7628..7662
                     /note="Rec2 site; pGREG recombinational cloning site II"
     misc_feature    7628..7633
                     /label=SalI restriction enzyme cut site
                     /note="SalI restriction enzyme cut site"
     regulatory      7632..7886
                     /regulatory_class="terminator"
                     /note="CYC1 terminator"
     terminator      7640..7887
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     misc_feature    7900..7933
                     /note="LoxP site (cre/lox KanMX marker rescue); Guldener, 
                     et. al. 1996, Nucleic Acids Research 24, 2519-2524"
     protein_bind    7900..7933
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     gene            complement(7991..9347)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     terminator      7991..8188
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(8194..9003)
                     /codon_start=1
                     /gene="aph(3)-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     misc_feature    8194..8939
                     /label=KanMX geneticin (G418) resistance marker
                     /note="KanMX geneticin (G418) resistance marker"
     promoter        complement(9004..9347)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     misc_feature    9402..9435
                     /note="LoxP site (cre/lox KanMX marker rescue); Guldener, 
                     et. al. 1996, Nucleic Acids Research 24, 2519-2524"
     protein_bind    9402..9435
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     promoter        complement(9472..9490)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(9500..9516)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"