LOCUS       40924_12650        4407 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Expression vector pColdI DNA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4407)
  AUTHORS   Qing G, Ma LC, Khorchid A, Swapna GV, Mal TK, Takayama MM, Xia B, 
            Phadtare S, Ke H, Acton T, Montelione GT, Ikura M, Inouye M.
  TITLE     Cold-shock induced high-yield protein production in Escherichia coli
  JOURNAL   Nat. Biotechnol. 22 (7), 877-882 (2004)
  PUBMED    15195104
REFERENCE   2  (bases 1 to 4407)
  AUTHORS   Takayama MM, Inouye M.
  TITLE     Direct Submission
  JOURNAL   Submitted (04-AUG-2004) Masanori Mitta Takayama, UMDNJ-Robert Wood 
            Johnson Medical School, Department of Biochemistry; 675 Hoes Lane, 
            Piscataway, NJ 08854-5635, USA (E-mail:mitsutam@takara-bio.co.jp, 
            Tel:81-77-543-7200, Fax:81-77-543-2494)
REFERENCE   3  (bases 1 to 4407)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 4407)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat. 
            Biotechnol."; date: "2004"; volume: "22"; issue: "7"; pages: 
            "877-882"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (04-AUG-2004) Masanori Mitta Takayama, UMDNJ-Robert Wood Johnson 
            Medical School, Department of Biochemistry"; volume: " 675 Hoes 
            Lane, Piscataway, NJ 08854-5635, USA 
            (E-mail:mitsutam@takara-bio.co.jp, Tel:81-77-543-7200, Fax"; pages: 
            "81-77-543-2494"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..4407
                     /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..300
                     /label=Factor Xa site
                     /note="Factor Xa recognition and cleavage site"
     misc_feature    301..360
                     /gene="cspA"
                     /note="MCS = Multiple cloning sites contain the follow 
                     restriction sites: NdeI, SacI, KpnI, XhoI, BamHI, EcoRI, 
                     HindIII, SalI, PstI, XbaI"
     3'UTR           368..512
                     /label=cspA 3'UTR
                     /note="3'UTR of the E. coli cold shock protein cspA gene
                     (Mitta et al., 1997)"
     rep_origin      complement(711..1166)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        1266..1370
                     /label=AmpR promoter
     CDS             1371..2228
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      2402..2990
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    complement(3128..3149)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(3165..4244)
                     /label=lacI
                     /note="lac repressor"
     promoter        complement(4245..4322)
                     /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 atcatcatat cgaaggtagg
      301 catatggagc tcggtaccct cgagggatcc gaattcaagc ttgtcgacct gcagtctaga
      361 taggtaatct ctgcttaaaa gcacagaatc taagatccct gccatttggc ggggattttt
      421 ttatttgttt tcaggaaata aataatcgat cgcgtaataa aatctattat tatttttgtg
      481 aagaataaat ttgggtgcaa tgagaatgcg caggcccttt cgtctcgcgc gtttcggtga
      541 tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc
      601 ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg
      661 ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata aaattgtaaa
      721 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
      781 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagcccgaga tagggttgag
      841 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
      901 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccca aatcaagttt
      961 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     1021 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     1081 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     1141 gcttaatgcg ccgctacagg gcgcgtacta tggttgcttt gacgtatgcg gtgtgaaata
     1201 ccgcacagat gcgtaaggag aaaataccgc atcaggcgtc aggtggcact tttcggggaa
     1261 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     1321 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     1381 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     1441 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     1501 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     1561 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     1621 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     1681 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     1741 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     1801 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     1861 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     1921 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     1981 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     2041 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     2101 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     2161 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     2221 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     2281 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     2341 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     2401 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     2461 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     2521 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     2581 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     2641 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     2701 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     2761 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     2821 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     2881 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     2941 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     3001 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacata gtcatgcccc
     3061 gcgcccaccg gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg
     3121 gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt
     3181 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     3241 tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg
     3301 cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc
     3361 aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta
     3421 tcgtcgtatc ccactaccga gatatccgca ccaacgcgca gcccggactc ggtaatggcg
     3481 cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc
     3541 tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt
     3601 tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga
     3661 cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg
     3721 accagatgct ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg
     3781 ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca
     3841 gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg
     3901 agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc
     3961 accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc
     4021 gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt
     4081 tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc
     4141 cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag
     4201 acaccggcat actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat
     4261 tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg
     4321 tccgggatct cgacgctctc ccttatgcga ctcctgcatt aggaagcagc ccagtagtag
     4381 gttgaggccg ttgagcaccg ccgccgc
//
