LOCUS       Exported               10154 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pAPG2, complete sequence.
ACCESSION   KJ814972
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10154)
  AUTHORS   Kang MK, Eom JH, Kim Y, Um Y, Woo HM.
  TITLE     Biosynthesis of pinene from glucose using metabolically-engineered 
            Corynebacterium glutamicum
  JOURNAL   Biotechnol. Lett. (2014) In press
  PUBMED    24930112
REFERENCE   2  (bases 1 to 10154)
  AUTHORS   Woo HM.
  TITLE     Direct Submission
  JOURNAL   Submitted (13-MAY-2014) Clean Energy Research Center, Korea 
            Institute of Science and Technology, Hwarangno 14-gil 5, 
            Seongbuk-gu, Seoul 136-791, Korea
REFERENCE   3  (bases 1 to 10154)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..10154
                     /organism="Expression vector pAPG2"
                     /host="Corynebacterium glutamicum"
                     /mol_type="other DNA"
                     /db_xref="taxon:1520899"
     regulatory      226..294
                     /regulatory_class="promoter"
                     /note="pTrc"
     promoter        266..294
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    302..318
                     /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)."
     CDS             370..2256
                     /codon_start=1
                     /product="pinene synthase"
                     /label=pinene synthase
                     /note="monoterpene synthase AgPS; codon-optimized for C. 
                     glutamicum"
                     /protein_id="AIF29188.1"
                     /translation="MALVSTAPLASKSCLHKSLISSTHELKALSRTIPALGMSRRGKSI
                     TPSISMSSTTVVTDDGVRRRMGDFHSNLWDDDVIQSLPTAYEEKSYLERAEKLIGEVKN
                     MFNSMSLEDGELMSPLNDLIQRLWIVDSLERLGIHRHFKDEIKSALDYVYSYWGENGIG
                     CGRESVVTDLNSTALGLRTLRLHGYPVSSDVFKAFKGQNGQFSCSENIQTDEEIRGVLN
                     LFRASLIAFPGEKIMDEAEIFSTKYLKEALQKIPVSSLSREIGDVLEYGWHTYLPRLEA
                     RNYIQVFGQDTENTKSYVKSKKLLELAKLEFNIFQSLQKRELESLVRWWKESGFPEMTF
                     CRHRHVEYYTLASCIAFEPQHSGFRLGFAKTCHLITVLDDMYDTFGTVDELELFTATMK
                     RWDPSSIDCLPEYMKGVYIAVYDTVNEMAREAEEAQGRDTLTYAREAWEAYIDSYMQEA
                     RWIATGYLPSFDEYYENGKVSCGHRISALQPILTMDIPFPDHILKEVDFPSKLNDLACA
                     ILRLRGDTRCYKADRARGEEASSISCYMKDNPGVSEEDALDHINAMISDVIKGLNWELL
                     KPDINVPISAKKHAFDIARAFHYGYKYRDGYSVANVETKSLVTRTLLESVPL"
     RBS             2263..2274
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2281..3426
                     /codon_start=1
                     /product="geranyl diphosphate synthase"
                     /label=geranyl diphosphate synthase
                     /note="AgGPPS2; codon-optimized for C. glutamicum"
                     /protein_id="AIF29189.1"
                     /translation="MAYSAMATMGYNGMAASCHTLHPTSPLKPFHGASTSLEAFNGEHM
                     GLLRGYSKRKLSSYKNPASRSSNATVAQLLNPPQKGKKAVEFDFNKYMDSKAMTVNEAL
                     NKAIPLRYPQKIYESMRYSLLAGGKRVRPVLCIAACELVGGTEELAIPTACAIEMIHTM
                     SLMHDDLPCIDNDDLRRGKPTNHKIFGEDTAVTAGNALHSYAFEHIAVSTSKTVGADRI
                     LRMVSELGRATGSEGVMGGQMVDIASEGDPSIDLQTLEWIHIHKTAMLLECSVVCGAII
                     GGASEIVIERARRYARCVGLLFQVVDDILDVTKSSDELGKTAGKDLISDKATYPKLMGL
                     EKAKEFSDELLNRAKGELSCFDPVKAAPLLGLADYVAFRQN"
     terminator      3653..3739
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3831..3858
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        3877..3968
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(7913..8728)
                     /codon_start=1
                     /gene="aph(3)-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIADALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      complement(9323..9868)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin."