LOCUS       Exported                8557 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pWCD2441, complete sequence.
ACCESSION   KR232291
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8557)
  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 8557)
  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 8557)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8557
                     /organism="Expression vector pWCD2441"
                     /mol_type="other DNA"
                     /db_xref="taxon:1655405"
     misc_feature    5..524
                     /label=similar to URA3 5
                     /note="similar to URA3 5"
     regulatory      573..1252
                     /regulatory_class="promoter"
     promoter        586..1252
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase"
     CDS             1259..2758
                     /codon_start=1
                     /product="CYP76AD1/P450 tyrosinase fusion protein"
                     /label=CYP76AD1/P450 tyrosinase fusion protein
                     /note="CYP76AD1 derived from Beta vulgaris - P450 
                     tyrosinase with W13L/F309L mutations"
                     /protein_id="AKH61456.1"
                     /translation="MDHATLAMILAILFISFHFIKLLFSQQTTKLLPPGPKPLPIIGNI
                     LEVGKKPHRSFANLAKIHGPLISLRLGSVTTIVVSSADVAKEMFLKKDHPLSNRTIPNS
                     VTAGDHHKLTMSWLPVSPKWRNFRKITAVHLLSPQRLDACQTFRHAKVQQLYEYVQECA
                     QKGQAVDIGKAAFTTSLNLLSKLFFSVELAHHKSHTSQEFKELIWNIMEDIGKPNYADY
                     FPILGCVDPSGIRRRLACSFDKLIAVFQGIICERLAPDSSTTTTTTTDDVLDVLLQLFK
                     QNELTMGEINHLLVDIFDAGTDTTSSTLEWVMTELIRNPEMMEKAQEEIKQVLGKDKQI
                     QESDIINLPYLQAIIKETLRLHPPTVFLLPRKADTDVELYGYIVPKDAQILVNLWAIGR
                     DPNAWQNADIFSPERFIGCEIDVKGRDFGLLPFGAGRRICPGMNLAIRMLTLMLATLLQ
                     FFNWKLEGDISPKDLDMDEKFGIALQKTKPLKLIPIPRYGS"
     misc_feature    1267
                     /label=9T>C relative to WT CYP76AD1
                     /note="9T>C relative to WT CYP76AD1"
     misc_feature    1295..1297
                     /label=W13L mutation relative to wild type CYP76AD1
                     /note="W13L mutation relative to wild type CYP76AD1"
     misc_feature    2183..2185
                     /label=F309L mutation relative to wildtype CYP76AD1
                     /note="F309L mutation relative to wildtype CYP76AD1"
     regulatory      2765..2988
                     /regulatory_class="terminator"
     regulatory      3092..3791
                     /regulatory_class="promoter"
     CDS             3798..4910
                     /codon_start=1
                     /product="ARO4 protein"
                     /label=ARO4 protein
                     /note="ARO4 derived from Saccharomyces cerevisiae with 
                     K229L feedback resistant mutation"
                     /protein_id="AKH61457.1"
                     /translation="MSESPMFAANGMPKVNQGAEEDVRILGYDPLASPALLQVQIPATP
                     TSLETAKRGRREAIDIITGKDDRVLVIVGPCSIHDLEAAQEYALRLKKLSDELKGDLSI
                     IMRAYLEKPRTTVGWKGLINDPDVNNTFNINKGLQSARQLFVNLTNIGLPIGSEMLDTI
                     SPQYLADLVSFGAIGARTTESQLHRELASGLSFPVGFKNGTDGTLNVAVDACQAAAHSH
                     HFMGVTLHGVAAITTTKGNEHCFVILRGGKKGTNYDAKSVAEAKAQLPAGSNGLMIDYS
                     HGNSNKDFRNQPKVNDVVCEQIANGENAITGVMIESNINEGNQGIPAEGKAGLKYGVSI
                     TDACIGWETTEDVLRKLAAAVRQRREVNKK"
     regulatory      4926..5150
                     /regulatory_class="terminator"
     regulatory      5199..5375
                     /regulatory_class="promoter"
     CDS             5376..6179
                     /codon_start=1
                     /product="URA3p uracil auxotrophy marker"
                     /label=URA3p uracil auxotrophy marker
                     /protein_id="AKH61459.1"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     regulatory      6180..6257
                     /regulatory_class="terminator"
     misc_feature    6262..6781
                     /label=similar to URA3 3
                     /note="similar to URA3 3"
     regulatory      complement(6794..6820)
                     /regulatory_class="terminator"
     CDS             complement(6821..7636)
                     /codon_start=1
                     /product="kanamycin resistance protein"
                     /label=kanamycin resistance protein
                     /protein_id="AKH61458.1"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     regulatory      complement(7637..7784)
                     /regulatory_class="promoter"
     rep_origin      complement(7785..8548)
                     /direction=LEFT
     rep_origin      7861..8449
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"