LOCUS       Exported                7596 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Shuttle vector pYT3, complete sequence.
ACCESSION   AF347016
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7596)
  AUTHORS   Tan X, Waterham HR, Veenhuis M, Cregg JM.
  TITLE     The Hansenula polymorpha PER8 gene encodes a novel peroxisomal 
            integral membrane protein involved in proliferation
  JOURNAL   J. Cell Biol. 128 (3), 307-319 (1995)
  PUBMED    7844145
REFERENCE   2  (bases 1 to 7596)
  AUTHORS   Baerends RJS., van Dijk R, Cregg JM, Veenhuis M.
  TITLE     Direct Submission
  JOURNAL   Submitted (08-FEB-2001) Eukaryotic Microbiology, Groningen 
            Biomolecular Sciences and Biotechnology Institute, Kerklaan 30, 
            Haren, Gn 9751 NN, the Netherlands
REFERENCE   3  (bases 1 to 7596)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7596
                     /organism="Shuttle vector pYT3"
                     /lab_host="Hansenula polymorpha strain CBS4732"
                     /mol_type="genomic DNA"
                     /note="Hansenula polymorpha-Escherichia coli shuttle 
                     vector; derived from pUC19"
                     /db_xref="taxon:154534"
     rep_origin      28..2724
                     /note="derived from Hansenula polymorpha; confers 
                     autonomous replication activity (HARS) in Hansenula 
                     polymorpha"
     RBS             1139..1147
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3 end of the 16S rRNA (Chen et al., 1994)"
     CDS             complement(3215..4309)
                     /codon_start=1
                     /gene="LEU2"
                     /product="3-isopropylmalate dehydrogenase"
                     /label=LEU2
                     /note="Leu2p; catalyzes the third step in the leucine 
                     biosynthesis pathway; derived from Saccharomyces 
                     cerevisiae; provides autonomous replication activity in 
                     Hansenula polymorpha"
                     /protein_id="AAK19746.1"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     gene            complement(3215..4309)
                     /gene="LEU2"
                     /label=LEU2
     promoter        complement(4310..4717)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(4989..5005)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5013..5029
                     /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)."
     promoter        complement(5037..5067)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5082..5103
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5391..5979)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6150..7010)
                     /codon_start=1
                     /gene="AmpR"
                     /product="beta-lactamase"
                     /label=AmpR
                     /protein_id="AAK19747.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(6150..7010)
                     /gene="AmpR"
                     /label=AmpR
     promoter        complement(7011..7115)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7589..9
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"