LOCUS       pCDF-2_Ek_LIC.        3561 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Bacterial vector with a CloDF13 (CDF) origin for 
            ligation-independent cloning (LIC) to express 6xHis- and S-tagged 
            proteins with an enterokinase site.
ACCESSION   .
VERSION     .
KEYWORDS    pCDF-2 Ek_LIC
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3561)
  AUTHORS   Novagen (EMD Millipore)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 3561)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     For LIC, linearize with BseRI and treat with T4 DNA polymerase plus 
            dTTP.
FEATURES             Location/Qualifiers
     source          1..3561
                     /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..73
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     CDS             77..94
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             98..112
                     /label=enterokinase site
                     /note="enterokinase recognition and cleavage site"
     CDS             146..190
                     /label=S-Tag
                     /note="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
     terminator      242..289
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(463..1251)
                     /label=SmR
                     /note="aminoglycoside adenylyltransferase (Murphy, 1985)"
     promoter        complement(1252..1343)
                     /label=AmpR promoter
     rep_origin      complement(1391..2129)
                     /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(2305..2326)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(2342..3421)
                     /label=lacI
                     /note="lac repressor"
     promoter        complement(3422..3499)
                     /label=lacI promoter
     promoter        3545..3561
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atggcacatc accaccacca tcacgtggat gacgacgaca agagtccggg
      121 cttctcctca aatctcgagt ctggtaaaga aaccgctgct gcgaaatttg aacgccagca
      181 catggactcg tctactagcg cagcttaatt aacctaggct gctgccaccg ctgagcaata
      241 actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaacctca
      301 ggcatttgag aagcacacgg tcacactgct tccggtagtc aataaaccgg taaaccagca
      361 atagacataa gcggctattt aacgaccctg ccctgaaccg acgaccgggt catcgtggcc
      421 ggatcttgcg gcccctcggc ttgaacgaat tgttagacat tatttgccga ctaccttggt
      481 gatctcgcct ttcacgtagt ggacaaattc ttccaactga tctgcgcgcg aggccaagcg
      541 atcttcttct tgtccaagat aagcctgtct agcttcaagt atgacgggct gatactgggc
      601 cggcaggcgc tccattgccc agtcggcagc gacatccttc ggcgcgattt tgccggttac
      661 tgcgctgtac caaatgcggg acaacgtaag cactacattt cgctcatcgc cagcccagtc
      721 gggcggcgag ttccatagcg ttaaggtttc atttagcgcc tcaaatagat cctgttcagg
      781 aaccggatca aagagttcct ccgccgctgg acctaccaag gcaacgctat gttctcttgc
      841 ttttgtcagc aagatagcca gatcaatgtc gatcgtggct ggctcgaaga tacctgcaag
      901 aatgtcattg cgctgccatt ctccaaattg cagttcgcgc ttagctggat aacgccacgg
      961 aatgatgtcg tcgtgcacaa caatggtgac ttctacagcg cggagaatct cgctctctcc
     1021 aggggaagcc gaagtttcca aaaggtcgtt gatcaaagct cgccgcgttg tttcatcaag
     1081 ccttacggtc accgtaacca gcaaatcaat atcactgtgt ggcttcaggc cgccatccac
     1141 tgcggagccg tacaaatgta cggccagcaa cgtcggttcg agatggcgct cgatgacgcc
     1201 aactacctct gatagttgag tcgatacttc ggcgatcacc gcttccctca tactcttcct
     1261 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     1321 atgtatttag aaaaataaac aaatagctag ctcactcggt cgctacgctc cgggcgtgag
     1381 actgcggcgg gcgctgcgga cacatacaaa gttacccaca gattccgtgg ataagcaggg
     1441 gactaacatg tgaggcaaaa cagcagggcc gcgccggtgg cgtttttcca taggctccgc
     1501 cctcctgcca gagttcacat aaacagacgc ttttccggtg catctgtggg agccgtgagg
     1561 ctcaaccatg aatctgacag tacgggcgaa acccgacagg acttaaagat ccccaccgtt
     1621 tccggcgggt cgctccctct tgcgctctcc tgttccgacc ctgccgttta ccggatacct
     1681 gttccgcctt tctcccttac gggaagtgtg gcgctttctc atagctcaca cactggtatc
     1741 tcggctcggt gtaggtcgtt cgctccaagc tgggctgtaa gcaagaactc cccgttcagc
     1801 ccgactgctg cgccttatcc ggtaactgtt cacttgagtc caacccggaa aagcacggta
     1861 aaacgccact ggcagcagcc attggtaact gggagttcgc agaggatttg tttagctaaa
     1921 cacgcggttg ctcttgaagt gtgcgccaaa gtccggctac actggaagga cagatttggt
     1981 tgctgtgctc tgcgaaagcc agttaccacg gttaagcagt tccccaactg acttaacctt
     2041 cgatcaaacc acctccccag gtggtttttt cgtttacagg gcaaaagatt acgcgcagaa
     2101 aaaaaggatc tcaagaagat cctttgatct tttctactga accgctctag atttcagtgc
     2161 aatttatctc ttcaaatgta gcacctgaag tcagccccat acgatataag ttgtaattct
     2221 catgttagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg
     2281 catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc
     2341 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     2401 cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga
     2461 cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca
     2521 cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac
     2581 atgagctgtc ttcggtatcg tcgtatccca ctaccgagat gtccgcacca acgcgcagcc
     2641 cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg
     2701 cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac
     2761 tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc
     2821 agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt
     2881 gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga
     2941 aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag
     3001 tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc
     3061 cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc
     3121 gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg
     3181 cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg
     3241 actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg
     3301 ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg
     3361 aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca
     3421 cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt
     3481 tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg
     3541 aaattaatac gactcactat a
//
