LOCUS       40924_9421        4955 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pCDF-RpALS, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4955)
  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 4955)
  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 4955)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 4955)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Biotechnol.
            Bioeng. (2018) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (15-JUN-2018) Dept. Chemical and Biomolecular Engineering, Korea 
            Advanced Institute of Science and Technology, Daehak-ro 291, Daejeon
            34141, South Korea"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..4955
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /label=lac operator
                     /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)."
     RBS             42..64
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             71..1246
                     /codon_start=1
                     /gene="ALS"
                     /product="aloesone synthase"
                     /label=ALS
                     /note="derived from Rheum palmatum"
                     /protein_id="AXN70001.1"
                     /translation="MADVLQEIRNSQKASGPATVLAIGTAHPPTCYPQADYPDFYFRVC
                     KSEHMTKLKKKMQFICDRSGIRQRFMFHTEENLGKNPGMCTFDGPSLNARQDMLIMEVP
                     KLGAEAAEKAIKEWGQDKSRITHLIFCTTTSNDMPGADYQFATLFGLNPGVSRTMVYQQ
                     GCFAGGTVLRLVKDIAENNKGARVLVVCSEIVAFAFRGPHEDHIDSLIGQLLFGDGAAA
                     LVVGTDIDESVERPIFQIMSATQATIPNSLHTMALHLTEAGLTFHLSKEVPKVVSDNME
                     ELMLEAFKPLGITDWNSIFWQVHPGGRAILDKIEEKLELTKDKMRDSRYILSEYGNLTS
                     ACVLFVMDEMRKRSFREGKQTTGDGYEWGVAIGLGPGLTVETVVLRSVPIP"
     gene            71..1246
                     /gene="ALS"
                     /label=ALS
     CDS             1257..1274
                     /label=6xHis
                     /note="6xHis affinity tag"
     promoter        1388..1406
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1407..1431
                     /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             1540..1584
                     /label=S-Tag
                     /note="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
     terminator      1636..1683
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1857..2645)
                     /label=SmR
                     /note="aminoglycoside adenylyltransferase (Murphy, 1985)"
     promoter        complement(2646..2737)
                     /label=AmpR promoter
     rep_origin      complement(2785..3523)
                     /direction=LEFT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     protein_bind    complement(3699..3720)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(3736..4815)
                     /label=lacI
                     /note="lac repressor"
     promoter        complement(4816..4893)
                     /label=lacI promoter
     regulatory      4939..4955
                     /label=T7 promoter
                     /note="T7 promoter"
                     /regulatory_class="promoter"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atggcagatg tcctgcagga gatccgcaac tcgcagaagg cgagcgggcc
      121 cgccacggtg ctcgccatcg gcactgccca tccaccgacg tgctaccctc aggccgacta
      181 ccccgacttc tacttccgag tttgcaagag cgagcacatg accaaactca agaagaaaat
      241 gcaattcatt tgtgacagat cggggataag gcagcggttt atgttccaca cggaagagaa
      301 cctggggaag aacccgggga tgtgcacatt cgacgggcca tcgctgaacg cgcggcagga
      361 catgctgatc atggaagtgc cgaagctggg ggcggaggcg gcggagaagg cgatcaagga
      421 gtgggggcag gacaagtccc ggatcaccca cctcatcttc tgcaccacca cgagcaacga
      481 catgcccggg gcggactacc agttcgccac cctgttcggg ctgaaccccg gcgtgagccg
      541 caccatggtc taccagcagg gctgcttcgc cgggggcacc gtgctgcgcc tggtcaagga
      601 catcgcggag aacaacaagg gggcgcgcgt gctggtggtg tgctcggaga tcgtggcctt
      661 cgccttccgc gggccccacg aggaccacat cgactccctc atcgggcagc tcctgttcgg
      721 ggacggggcc gccgccctcg tggtcgggac agacatcgac gagagcgtcg agaggcccat
      781 cttccagatc atgtcggcga cccaggcgac catccccaac tcgctgcaca ccatggctct
      841 ccatctgacg gaggcggggc tgaccttcca tctcagcaag gaggtgccca aggtggtgag
      901 cgacaacatg gaggagctca tgctcgaggc cttcaagccg ctcgggataa ccgattggaa
      961 ctccatattc tggcaagtgc atcccggggg tagagccatc cttgacaaga tcgaggagaa
     1021 gctggagctc accaaggata agatgcggga ttcccgctac atcttgagcg agtacgggaa
     1081 tctcaccagc gcctgtgtgc tctttgtcat ggacgagatg aggaagaggt ccttccggga
     1141 agggaagcag accaccggag acggctacga gtggggtgtc gccatcggat tggggcccgg
     1201 tcttaccgtc gagaccgttg tcttgcgtag cgtccccatt ccctaatggc agcagccatc
     1261 accatcatca ccacagccag gatccgaatt cgagctcggc gcgcctgcag gtcgacaagc
     1321 ttgcggccgc ataatgctta agtcgaacag aaagtaatcg tattgtacac ggccgcataa
     1381 tcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc cccatcttag
     1441 tatattagtt aagtataaga aggagatata catatggcag atctcaattg gatatcggcc
     1501 ggccacgcga tcgctgacgt cggtaccctc gagtctggta aagaaaccgc tgctgcgaaa
     1561 tttgaacgcc agcacatgga ctcgtctact agcgcagctt aattaaccta ggctgctgcc
     1621 accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt
     1681 ttgctgaaac ctcaggcatt tgagaagcac acggtcacac tgcttccggt agtcaataaa
     1741 ccggtaaacc agcaatagac ataagcggct atttaacgac cctgccctga accgacgacc
     1801 gggtcatcgt ggccggatct tgcggcccct cggcttgaac gaattgttag acattatttg
     1861 ccgactacct tggtgatctc gcctttcacg tagtggacaa attcttccaa ctgatctgcg
     1921 cgcgaggcca agcgatcttc ttcttgtcca agataagcct gtctagcttc aagtatgacg
     1981 ggctgatact gggccggcag gcgctccatt gcccagtcgg cagcgacatc cttcggcgcg
     2041 attttgccgg ttactgcgct gtaccaaatg cgggacaacg taagcactac atttcgctca
     2101 tcgccagccc agtcgggcgg cgagttccat agcgttaagg tttcatttag cgcctcaaat
     2161 agatcctgtt caggaaccgg atcaaagagt tcctccgccg ctggacctac caaggcaacg
     2221 ctatgttctc ttgcttttgt cagcaagata gccagatcaa tgtcgatcgt ggctggctcg
     2281 aagatacctg caagaatgtc attgcgctgc cattctccaa attgcagttc gcgcttagct
     2341 ggataacgcc acggaatgat gtcgtcgtgc acaacaatgg tgacttctac agcgcggaga
     2401 atctcgctct ctccagggga agccgaagtt tccaaaaggt cgttgatcaa agctcgccgc
     2461 gttgtttcat caagccttac ggtcaccgta accagcaaat caatatcact gtgtggcttc
     2521 aggccgccat ccactgcgga gccgtacaaa tgtacggcca gcaacgtcgg ttcgagatgg
     2581 cgctcgatga cgccaactac ctctgatagt tgagtcgata cttcggcgat caccgcttcc
     2641 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     2701 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag ctagctcact cggtcgctac
     2761 gctccgggcg tgagactgcg gcgggcgctg cggacacata caaagttacc cacagattcc
     2821 gtggataagc aggggactaa catgtgaggc aaaacagcag ggccgcgccg gtggcgtttt
     2881 tccataggct ccgccctcct gccagagttc acataaacag acgcttttcc ggtgcatctg
     2941 tgggagccgt gaggctcaac catgaatctg acagtacggg cgaaacccga caggacttaa
     3001 agatccccac cgtttccggc gggtcgctcc ctcttgcgct ctcctgttcc gaccctgccg
     3061 tttaccggat acctgttccg cctttctccc ttacgggaag tgtggcgctt tctcatagct
     3121 cacacactgg tatctcggct cggtgtaggt cgttcgctcc aagctgggct gtaagcaaga
     3181 actccccgtt cagcccgact gctgcgcctt atccggtaac tgttcacttg agtccaaccc
     3241 ggaaaagcac ggtaaaacgc cactggcagc agccattggt aactgggagt tcgcagagga
     3301 tttgtttagc taaacacgcg gttgctcttg aagtgtgcgc caaagtccgg ctacactgga
     3361 aggacagatt tggttgctgt gctctgcgaa agccagttac cacggttaag cagttcccca
     3421 actgacttaa ccttcgatca aaccacctcc ccaggtggtt ttttcgttta cagggcaaaa
     3481 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta ctgaaccgct
     3541 ctagatttca gtgcaattta tctcttcaaa tgtagcacct gaagtcagcc ccatacgata
     3601 taagttgtaa ttctcatgtt agtcatgccc cgcgcccacc ggaaggagct gactgggttg
     3661 aaggctctca agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta
     3721 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     3781 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct
     3841 tttcaccagt gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg
     3901 cagcaagcgg tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa
     3961 cggcgggata taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatgtccgc
     4021 accaacgcgc agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt
     4081 ggcaaccagc atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa
     4141 accggacatg gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt
     4201 gagatattta tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc
     4261 taacagcgcg atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc
     4321 gtcttcatgg gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa
     4381 cgccggaaca ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata
     4441 gttaatgatc agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc
     4501 ttcgacgccg cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg
     4561 agatttaatc gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac
     4621 gccaatcagc aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt
     4681 cagctccgcc atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg
     4741 gttcaccacg cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa
     4801 cgttactggt ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat
     4861 accgcgaaag gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg
     4921 actcctgcat taggaaatta atacgactca ctata
//
