LOCUS       Exported               12051 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pWCD2353, complete sequence.
ACCESSION   KR232300
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12051)
  AUTHORS   DeLoache WC, Russ ZN, Narcross L, Gonzales AM, Martin VJ, Dueber JE.
  TITLE     An enzyme-coupled biosensor enables (S)-reticuline production in 
            yeast from glucose
  JOURNAL   Nat. Chem. Biol. 11 (7), 465-471 (2015)
  PUBMED    25984720
REFERENCE   2  (bases 1 to 12051)
  AUTHORS   DeLoache WC, Russ ZN, Narcross L, Gonzales AM, Martin VJJ., Dueber 
            JE.
  TITLE     Direct Submission
  JOURNAL   Submitted (22-APR-2015) Department of Bioengineering, University of 
            California, Berkeley, 2151 Berkeley Way Room 542, Berkeley, CA 
            94704, USA
REFERENCE   3  (bases 1 to 12051)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..12051
                     /organism="Expression vector pWCD2353"
                     /mol_type="other DNA"
                     /db_xref="taxon:1655396"
     regulatory      57..736
                     /regulatory_class="promoter"
     promoter        70..736
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase"
     CDS             743..1789
                     /codon_start=1
                     /product="6-O-methyltransferase"
                     /label=6-O-methyltransferase
                     /note="derived from Papaver somniferum"
                     /protein_id="AKH61490.1"
                     /translation="METVSKIDQQNQAKIWKQIYGFAESLVLKCAVQLEIAETLHNNVK
                     PMSLSELASKLPVAQPVNEDRLFRIMRYLVHMELFKIDATTQKYSLAPPAKYLLRGWEK
                     SMVDSILCINDKDFLAPWHHLGDGLTGNCDAFEKALGKSIWVYMSVNPEKNQLFNAAMA
                     CDTRLVTSALANECKSIFSDGISTLVDVGGGTGTAVKAISKAFPDIKCTIYDLPHVIAD
                     SPEIPNITKISGDMFKSIPSADAIFMKCILHDWNDDECIQILKRCKEALPKGGKVIIVD
                     VVIDMDSTHPYAKIRLTLDLDMMLNTGGKERTKEEWKTLFDAAGFASHKVTQISAVQSV
                     IEAYPYGS"
     regulatory      1796..2020
                     /regulatory_class="terminator"
     terminator      1796..1983
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     regulatory      2054..2753
                     /regulatory_class="promoter"
     CDS             2760..3821
                     /codon_start=1
                     /product="N-methyltransferase"
                     /label=N-methyltransferase
                     /note="derived from Papaver somniferum"
                     /protein_id="AKH61489.1"
                     /translation="MQLKAKEELLRNMELGLIPDQEIRQLIRVELEKRLQWGYKETHEE
                     QLSQLLDLVHSLKGMKMATEMENLDLKLYEAPMEFLKIQHGSNMKQSAGYYTDESTTLD
                     EAEIAMLDLYMERAQIKDGQSVLDLGCGLGAVALFGANKFKKCQFTGVTSSVEQKDYIE
                     GKCKELKLTNVKVLLADITTYETEERFDRIFAVELIEHMKNYQLLLKKISEWMKDDGLL
                     FVEHVCHKTLAYHYEPVDAEDWYTNYIFPAGTLTLSSASMLLYFQDDVSVVNQWTLSGK
                     HYSRSHEEWLKNMDKNIVEFKEIMRSITKTEKEAIKLLNFWRIFCMCGAELFGYKNGEE
                     WMLTHLLFKKKGS"
     regulatory      3828..4052
                     /regulatory_class="terminator"
     regulatory      4086..4785
                     /regulatory_class="promoter"
     CDS             4792..5871
                     /codon_start=1
                     /product="4 O-methyltransferase"
                     /label=4 O-methyltransferase
                     /note="derived from Papaver somniferum"
                     /protein_id="AKH61488.1"
                     /translation="MGSLDAKPAAATQEVSIKDQAQLWNIIYGFADSLVLRCAVEIGIA
                     DIIKNNDGAITLAQLAAKLPITNVSSDYLYRMVRYLVHLNIIEQETCNGGVEKVYSLKP
                     VGTLLLRDAERSMVPMILGMTQKDFMVSWHFMKEGLGNGSTTAFEKGMGMDIWKYLEGN
                     PDQSQLFNEGMAGETRLLTKTLIEDCRDTFQGLDSLVDIGGGNGTTIKAIYEAFPHIKC
                     TLYDLPHVVANSHDLPNIEKVPGDMFKSVPSAQAILLKLILHDWTDEECVNILKKCKEA
                     IPKETGKVIIVDVALEEESNHELTKTRLILDIDMLVNTGGRERTADDWENLLKRAGFRS
                     HKIRPIRAIQSVIEAFPGS"
     regulatory      5878..6102
                     /regulatory_class="terminator"
     regulatory      6206..6905
                     /regulatory_class="promoter"
     promoter        6485..6892
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             6912..8384
                     /codon_start=1
                     /product="NMCH CYP80B1 P450 N-methylcoclaurine hydroxylase 
                     from Eschscholzia californica"
                     /label=NMCH CYP80B1 P450 N-methylcoclaurine hydroxylas
                     /protein_id="AKH61487.1"
                     /translation="MEVVTVALIAVIISSILYLLFGSSGHKNLPPGPKPWPIVGNLLQL
                     GEKPHAQFAELAQTYGDIFTLKMGTETVVVASTSSAASEILKTHDRILSARYVFQSFRV
                     KGHVENSIVWSDCTETWKNLRKVCRTELFTQKMIESQAHVREKKCEEMVEYLMKKQGEE
                     VKIVEVIFGTLVNIFGNLIFSQNIFELGDPNSGSSEFKEYLWRMLELGNSTNPADYFPM
                     LGKFDLFGQRKEVAECLKGIYAIWGAMLQERKLAKKVDGYQSKNDFVDVCLDSGLNDYQ
                     INALLMELFGAGTETSASTIEWAMTELTKNPKITAKLRSELQTVVGERSVKESDFPNLP
                     YLEATVKETLRLHPPTPLLLPRRALETCTILNYTIPKDCQIMVNAWGIGRDPKTWIDPL
                     TFSPERFLNSSVDFRGNDFSLIPFGAGRRICPGLPIANQFIALLVATFVQNLDWCLPNG
                     MSVDHLIVEEKFGLTLQKEPPLFIVPKSRVGS"
     regulatory      8391..8614
                     /regulatory_class="terminator"
     regulatory      8663..8899
                     /regulatory_class="promoter"
     promoter        8712..8899
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             8900..9562
                     /codon_start=1
                     /product="HIS3 histidine auxotrophy marker"
                     /label=HIS3 histidine auxotrophy marker
                     /protein_id="AKH61492.1"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     regulatory      9563..9772
                     /regulatory_class="terminator"
     misc_feature    9777..10280
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     rep_origin      9777..9893
     rep_origin      9894..10284
     regulatory      complement(10297..10323)
                     /regulatory_class="terminator"
     CDS             complement(10324..11139)
                     /codon_start=1
                     /product="kanamycin resistance protein"
                     /label=kanamycin resistance protein
                     /protein_id="AKH61491.1"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     regulatory      complement(11140..11287)
                     /regulatory_class="promoter"
     rep_origin      complement(11288..12051)
                     /direction=LEFT
     rep_origin      11364..11952
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"