LOCUS       40924_26146        4366 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pJJDuet30, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4366)
  AUTHORS   Zuger S, Iwai H.
  TITLE     Intein-based biosynthetic incorporation of unlabeled protein tags 
            into isotopically labeled proteins for NMR studies
  JOURNAL   Nat. Biotechnol. 23 (6), 736-740 (2005)
  PUBMED    15908942
REFERENCE   2  (bases 1 to 4366)
  AUTHORS   Zueger S, Iwai H.
  TITLE     Direct Submission
  JOURNAL   Submitted (22-APR-2005) Chemistry, University of Saskatchewan, 110 
            Science Place, Saskatoon, SK S7N 5C9, Canada
REFERENCE   3  (bases 1 to 4366)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 4366)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat. 
            Biotechnol."; date: "2005"; volume: "23"; issue: "6"; pages: 
            "736-740"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (22-APR-2005) Chemistry, University of Saskatchewan, 110 Science 
            Place, Saskatoon, SK S7N 5C9, Canada"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..4366
                     /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             83..100
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     CDS             110..127
                     /codon_start=1
                     /label=thrombin site
                     /note="thrombin recognition and cleavage site"
                     /translation="LVPRGS"
     CDS             137..298
                     /codon_start=1
                     /label=GB1
                     /note="B1 domain of Streptococcal protein G"
                     /translation="YKLILNGKTLKGETTTEAVDAATAEKVFKQYANDNGVDGEWTYDD
                     ATKTYTVTE"
     promoter        751..769
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    770..794
                     /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             903..947
                     /codon_start=1
                     /label=S-Tag
                     /note="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /translation="KETAAAKFERQHMDS"
     terminator      999..1046
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1279..2091)
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     promoter        complement(2092..2183)
                     /label=AmpR promoter
     rep_origin      complement(2199..2948)
                     /direction=LEFT
                     /label=RSF ori
                     /note="Plasmids containing the RSF 1030 origin of
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     protein_bind    complement(3110..3131)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(3147..4226)
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(4227..4304)
                     /label=lacI promoter
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atgggcagca gccatcatca tcatcatcac agcagcggcc tggtgccgcg
      121 cggcagccat atgcagtaca aacttatcct gaacggtaaa accctgaaag gtgaaaccac
      181 caccgaagct gttgacgctg ctaccgcgga aaaagttttc aaacagtacg ctaacgacaa
      241 cggtgttgac ggtgaatgga cctacgacga cgctaccaaa acctacaccg taacggaagg
      301 atcctgcctc agttttggta ccgaaatttt aaccgttgag tacggcccat tgcccattgg
      361 caaaattgtg agtgaagaaa ttaattgttc tgtgtacagt gttgatccag aagggagagt
      421 ttacacccag gcgatcgccc aatggcatga ccggggagag caggaagtat tggaatatga
      481 attggaagat ggttcagtaa tccgagctac ctctgaccac cgctttttaa ccaccgatta
      541 tcaactgttg gcgatcgaag aaatttttgc taggcaactg gacttgttga ctttagaaaa
      601 tattaagcaa actgaagaag ctcttgacaa ccatcgtctt ccctttccat tacttgacgc
      661 aggtacaatt aaataaataa agcttgcggc cgcataatgc ttaagtcgaa cagaaagtaa
      721 tcgtattgta cacggccgca taatcgaaat taatacgact cactataggg gaattgtgag
      781 cggataacaa ttccccatct tagtatatta gttaagtata agaaggagat atacatatgg
      841 cagatctcaa ttggatatcg gccggccacg cgatcgctga cgtcggtacc ctcgagtctg
      901 gtaaagaaac cgctgctgcg aaatttgaac gccagcacat ggactcgtct actagcgcag
      961 cttaattaac ctaggctgct gccaccgctg agcaataact agcataaccc cttggggcct
     1021 ctaaacgggt cttgaggggt tttttgctga aacctcaggc atttgagaag cacacggtca
     1081 cactgcttcc ggtagtcaat aaaccggtaa accagcaata gacataagcg gctatttaac
     1141 gaccctgccc tgaaccgacg acaagctgac gaccgggtct ccgcaagtgg cacttttcgg
     1201 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     1261 ctcatgaatt aattcttaga aaaactcatc gagcatcaaa tgaaactgca atttattcat
     1321 atcaggatta tcaataccat atttttgaaa aagccgtttc tgtaatgaag gagaaaactc
     1381 accgaggcag ttccatagga tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc
     1441 aacatcaata caacctatta atttcccctc gtcaaaaata aggttatcaa gtgagaaatc
     1501 accatgagtg acgactgaat ccggtgagaa tggcaaaagt ttatgcattt ctttccagac
     1561 ttgttcaaca ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt
     1621 attcattcgt gattgcgcct gagcgagacg aaatacgcgg tcgctgttaa aaggacaatt
     1681 acaaacagga atcgaatgca accggcgcag gaacactgcc agcgcatcaa caatattttc
     1741 acctgaatca ggatattctt ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt
     1801 gagtaaccat gcatcatcag gagtacggat aaaatgcttg atggtcggaa gaggcataaa
     1861 ttccgtcagc cagtttagtc tgaccatctc atctgtaaca tcattggcaa cgctaccttt
     1921 gccatgtttc agaaacaact ctggcgcatc gggcttccca tacaatcgat agattgtcgc
     1981 acctgattgc ccgacattat cgcgagccca tttataccca tataaatcag catccatgtt
     2041 ggaatttaat cgcggcctag agcaagacgt ttcccgttga atatggctca tactcttcct
     2101 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     2161 atgtatttag aaaaataaac aaataggcat gcagcgctct tccgcttcct cgctcactga
     2221 ctcgctacgc tcggtcgttc gactgcggcg agcggtgtca gctcactcaa aagcggtaat
     2281 acggttatcc acagaatcag gggataaagc cggaaagaac atgtgagcaa aaagcaaagc
     2341 accggaagaa gccaacgccg caggcgtttt tccataggct ccgcccccct gacgagcatc
     2401 acaaaaatcg acgctcaagc cagaggtggc gaaacccgac aggactataa agataccagg
     2461 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     2521 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgttggt
     2581 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     2641 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     2701 acttatcgcc actggcagca gccattggta actgatttag aggactttgt cttgaagtta
     2761 tgcacctgtt aaggctaaac tgaaagaaca gattttggtg agtgcggtcc tccaacccac
     2821 ttaccttggt tcaaagagtt ggtagctcag cgaaccttga gaaaaccacc gttggtagcg
     2881 gtggtttttc tttatttatg agatgatgaa tcaatcggtc tatcaagtca acgaacagct
     2941 attccgttac tctagatttc agtgcaattt atctcttcaa atgtagcacc tgaagtcagc
     3001 cccatacgat ataagttgta attctcatgt tagtcatgcc ccgcgcccac cggaaggagc
     3061 tgactgggtt gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct
     3121 aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     3181 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg
     3241 gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc
     3301 tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg
     3361 atggtggtta acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc
     3421 gagatgtccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc
     3481 atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg
     3541 gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt
     3601 tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt
     3661 aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc
     3721 agtcgcgtac cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca
     3781 tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca
     3841 tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc
     3901 gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt
     3961 tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg
     4021 gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg
     4081 ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg
     4141 tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg
     4201 acatcgtata acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc
     4261 tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc
     4321 tcccttatgc gactcctgca ttaggaaatt aatacgactc actata
//
