LOCUS       40924_12645        6090 bp DNA     circular SYN 25-JUL-2018
DEFINITION  Vector for cold shock-induced expression in E. coli of a protein 
            fused to the Trigger Factor chaperone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6090)
  AUTHORS   TaKaRa
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 6090)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6090
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        15..81
                     /label=cspA promoter
                     /note="promoter of the E. coli cold shock protein cspA gene
                     (Mitta et al., 1997)"
     protein_bind    84..100
                     /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)."
     5'UTR           122..255
                     /label=cspA 5'UTR
                     /note="5'UTR of the E. coli cold shock protein cspA gene
                     (Mitta et al., 1997)"
     CDS             256..270
                     /label=TEE
                     /note="translation enhancing element for E. coli (Qing et
                     al., 2004)"
     CDS             271..288
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             289..1584
                     /label=Trigger Factor
                     /note="ribosome-associated chaperone from E. coli (Hoffmann
                     et al., 2010)"
     CDS             1594..1617
                     /label=HRV 3C site
                     /note="recognition and cleavage site for human rhinovirus
                     3C and PreScission proteases"
     CDS             1627..1644
                     /label=thrombin site
                     /note="thrombin recognition and cleavage site"
     CDS             1651..1662
                     /label=Factor Xa site
                     /note="Factor Xa recognition and cleavage site"
     CDS             1666..1998
                     /gene="mazF"
                     /label=mazF
                     /note="Endoribonuclease toxin MazF from Escherichia coli
                     (strain K12). Accession#: P0AE70"
     3'UTR           2051..2195
                     /label=cspA 3'UTR
                     /note="3'UTR of the E. coli cold shock protein cspA gene
                     (Mitta et al., 1997)"
     rep_origin      complement(2394..2849)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2949..3053
                     /label=AmpR promoter
     CDS             3054..3911
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      4085..4673
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    complement(4811..4832)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(4848..5927)
                     /label=lacI
                     /note="lac repressor"
     promoter        complement(5928..6005)
                     /label=lacI promoter
ORIGIN
        1 aaggaatggt gtggccgatt aatcataaat atgaaaaata attgttgcat cacccgccaa
       61 tgcgtggctt aatgcacatc aaattgtgag cggataacaa tttgatgtgc tagcgcatat
      121 ccagtgtagt aaggcaagtc ccttcaagag ttatcgttga tacccctcgt agtgcacatt
      181 cctttaacgc ttcaaaatct gtaaagcacg ccatatcgcc gaaaggcaca cttaattatt
      241 aagaggtaat acaccatgaa tcacaaagtg catcatcatc atcatcacat gcaagtttca
      301 gttgaaacca ctcaaggcct tggccgccgt gtaacgatta ctatcgctgc tgacagcatc
      361 gagaccgctg ttaaaagcga gctggtcaac gttgcgaaaa aagtacgtat tgacggcttc
      421 cgcaagggca aagtgccaat gaatatcgtt gctcagcgtt atggcgcgtc tgtacgccag
      481 gacgttctgg gtgacctgat gagccgtaac ttcattgacg ccatcattaa agaaaaaatc
      541 aatccggctg gcgcaccgac ttatgttccg ggcgaataca agctgggtga agacttcact
      601 tactctgtag agtttgaagt ttatccggaa gttgaactgc aaggtctgga agcgatcgaa
      661 gttgaaaaac cgatcgttga agtgaccgac gctgacgttg acggcatgct ggatactctg
      721 cgtaaacagc aggcgacctg gaaagaaaaa gacggcgctg ttgaagcaga agaccgcgtg
      781 accatcgact tcaccggttc tgtagacggc gaagagttcg aaggcggtaa agcgtctgat
      841 ttcgtactgg cgatgggcca gggtcgtatg atcccgggct ttgaagacgg tatcaaaggc
      901 cacaaagctg gcgaagagtt caccatcgac gtgaccttcc cggaagaata ccacgcagaa
      961 aacctgaaag gtaaagcagc gaaattcgct atcaacctga agaaagttga agagcgtgaa
     1021 ctgccggaac tgaccgcaga gttcatcaaa cgtttcggcg ttgaagatgg ttccgtagaa
     1081 ggtctgcgcg ctgaagtgcg taaaaacatg gagcgcgagc tgaagagcgc catccgtaac
     1141 cgcgttaagt ctcaggcgat cgaaggtctg gtaaaagcta acgacatcga cgtaccggct
     1201 gcgctgatcg acagcgaaat cgacgttctg cgtcgccagg ctgcacagcg tttcggtggc
     1261 aacgaaaaac aagctctgga actgccgcgc gaactgttcg aagaacaggc taaacgccgc
     1321 gtagttgttg gcctgctgct gggcgaagtt atccgcacca acgagctgaa agctgacgaa
     1381 gagcgcgtga aaggcctgat cgaagagatg gcttctgcgt acgaagatcc gaaagaagtt
     1441 atcgagttct acagcaaaaa caaagaactg atggacaaca tgcgcaatgt tgctctggaa
     1501 gaacaggctg ttgaagctgt actggcgaaa gcgaaagtga ctgaaaaaga aaccactttc
     1561 aacgagctga tgaaccagca ggcgtccgcg ggtctggaag ttctgttcca ggggccctcc
     1621 gcgggtctgg tgccacgcgg tagtggtggt atcgaaggta ggcatatggt aagccgatac
     1681 gtacccgata tgggcgatct gatttgggtt gattttgacc cgacaaaagg tagcgagcaa
     1741 gctggacatc gtccagctgt tgtcctgagt cctttcatgt acaacaacaa aacaggtatg
     1801 tgtctgtgtg ttccttgtac aacgcaatca gctggatatc cgttcgaagt tgttttatcc
     1861 ggtcaggaac gtgatggcgt agcgttagct gatcaggtaa aaagtatcgc ctggcgggca
     1921 agaggagcaa cgaagaaagg aacagttgcc ccagaggaat tacaactcat taaagccaaa
     1981 attaacgtac tgattgggta gctcgaggga tccgaattca agcttgtcga cctgcagtct
     2041 agataggtaa tctctgctta aaagcacaga atctaagatc cctgccattt ggcggggatt
     2101 tttttatttg ttttcaggaa ataaataatc gatcgcgtaa taaaatctat tattattttt
     2161 gtgaagaata aatttgggtg caatgagaat gcgcaggccc tttcgtctcg cgcgtttcgg
     2221 tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta
     2281 agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg
     2341 gggctggctt aactatgcgg catcagagca gattgtactg agagtgcacc ataaaattgt
     2401 aaacgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa
     2461 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagcccg agatagggtt
     2521 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa
     2581 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac ccaaatcaag
     2641 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
     2701 tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
     2761 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
     2821 cgcgcttaat gcgccgctac agggcgcgta ctatggttgc tttgacgtat gcggtgtgaa
     2881 ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gtcaggtggc acttttcggg
     2941 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     3001 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     3061 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     3121 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     3181 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     3241 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     3301 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     3361 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     3421 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     3481 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     3541 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     3601 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     3661 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     3721 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     3781 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     3841 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     3901 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     3961 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     4021 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     4081 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     4141 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     4201 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     4261 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     4321 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     4381 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     4441 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     4501 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     4561 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     4621 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     4681 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atagtcatgc
     4741 cccgcgccca ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc
     4801 ccggtgccta atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc
     4861 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     4921 gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga
     4981 ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc
     5041 agcaggcgaa aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg
     5101 gtatcgtcgt atcccactac cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg
     5161 gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg
     5221 ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc
     5281 cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc
     5341 agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat
     5401 gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg
     5461 atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc
     5521 acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc
     5581 gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac
     5641 accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac
     5701 ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc
     5761 agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt
     5821 tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa
     5881 gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg
     5941 aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg
     6001 gtgtccggga tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag
     6061 taggttgagg ccgttgagca ccgccgccgc
//
