LOCUS       Exported                6881 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pTacCDF-VvSTS-At4CL1m, complete sequence.
ACCESSION   MH488945
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6881)
  AUTHORS   Kim B, Binkley R, Kim HU, Lee SY.
  TITLE     Metabolic engineering of Escherichia coli for the enhanced 
            production of l-tyrosine
  JOURNAL   Biotechnol. Bioeng. (2018) In press
  PUBMED    30019750
REFERENCE   2  (bases 1 to 6881)
  AUTHORS   Yang D, Kim WJ, Yoo SM, Choi JH, Ha SH, Lee MH, Lee SY.
  TITLE     Direct Submission
  JOURNAL   Submitted (15-JUN-2018) Dept. Chemical and Biomolecular Engineering,
            Korea Advanced Institute of Science and Technology, Daehak-ro 291, 
            Daejeon 34141, South Korea
REFERENCE   3  (bases 1 to 6881)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6881
                     /organism="Cloning vector pTacCDF-VvSTS-At4CL1m"
                     /lab_host="Escherichia coli"
                     /mol_type="other DNA"
                     /db_xref="taxon:2301170"
     regulatory      195..223
                     /regulatory_class="promoter"
                     /note="Tac promoter"
     promoter        195..223
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    231..247
                     /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             269..1447
                     /codon_start=1
                     /gene="STS"
                     /product="stilbene synthase"
                     /label=STS
                     /note="Vitis vinifera"
                     /protein_id="AXN70055.1"
                     /translation="MASVEEFRNAQRAKGPATILAIGTATPDHCVYQSDYADFYFRVTK
                     SEHMTALKKKFNRICDKSMIKKRYIHLTEEMLEEHPNIGAYMAPSLNIRQEIITAEVPK
                     LGKEAALKALKEWGQPKSKITHLVFCTTSGVEMPGADYKLANLLGLEPSVRRVMLYHQG
                     CYAGGTVLRTAKDLAENNAGARVLVVCSEITVVTFRGPSEDALDSLVGQALFGDGSAAV
                     IVGSDPDISIERPLFQLVSAAQTFIPNSAGAIAGNLREVGLTFHLWPNVPTLISENIEK
                     CLTQAFDPLGISDWNSLFWIAHPGGPAILDAVEAKLNLDKKKLEATRHVLSEYGNMSSA
                     CVLFILDEMRKKSLKGERATTGEGLDWGVLFGFGPGLTIETVVLHSIPMVTN"
     gene            269..1447
                     /gene="STS"
                     /label=STS
     regulatory      1656..1684
                     /regulatory_class="promoter"
                     /note="Tac promoter"
     promoter        1656..1684
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1692..1708
                     /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             1730..3415
                     /codon_start=1
                     /gene="4CL1m"
                     /product="4-coumarate:CoA ligase 1 (4AT mutant)"
                     /label=4CL1m
                     /note="I250L/N404K/I461V; derived from Arabidopsis 
                     thaliana"
                     /protein_id="AXN70056.1"
                     /translation="MAPQEQAVSQVMEKQSNNNNSDVIFRSKLPDIYIPNHLSLHDYIF
                     QNISEFATKPCLINGPTGHVYTYSDVHVISRQIAANFHKLGVNQNDVVMLLLPNCPEFV
                     LSFLAASFRGATATAANPFFTPAEIAKQAKASNTKLIITEARYVDKIKPLQNDDGVVIV
                     CIDDNESVPIPEGCLRFTELTQSTTEASEVIDSVEISPDDVVALPYSSGTTGLPKGVML
                     THKGLVTSVAQQVDGENPNLYFHSDDVLLCVLPMFHIYALNSIMLCGLRVGAAILIMPK
                     FEINLLLELIQRCKVTVAPMVPPIVLAIAKSSETEKYDLSSIRVVKSGAAPLGKELEDA
                     VNAKFPNAKLGQGYGMTEAGPVLAMSLGFAKEPFPVKSGACGTVVRNAEMKIVDPDTGD
                     SLSRKQPGEICIRGHQIMKGYLNNPAATAETIDKDGWLHTGDIGLIDDDDELFIVDRLK
                     ELVKYKGFQVAPAELEALLIGHPDITDVAVVAMKEEAAGEVPVAFVVKSKDSELSEDDV
                     KQFVSKQVVFYKRINKVFFTESIPKAPSGKILRKDLRAKLANGL"
     gene            1730..3415
                     /gene="4CL1m"
                     /label=4CL1m
     regulatory      3459..3604
                     /regulatory_class="terminator"
                     /note="rrnBT1T2"
     CDS             complement(3797..4588)
                     /codon_start=1
                     /product="spectinomycin resistance protein"
                     /label=spectinomycin resistance protein
                     /protein_id="AXN70057.1"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        complement(4589..4680)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4728..5466)
                     /direction=LEFT
                     /label=CDF
                     /note="CDF"
     CDS             complement(5676..6758)
                     /codon_start=1
                     /gene="lacIq"
                     /product="lac repressor protein"
                     /label=lacIq
                     /protein_id="AXN70058.1"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     gene            complement(5676..6758)
                     /gene="lacIq"
                     /label=lacIq
     promoter        complement(6759..6836)
                     /gene="lacI"
                     /label=lacI promoter