LOCUS       pRSF-2_Ek_LIC.        3609 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Bacterial vector with an RSF 1030 origin for ligation-independent 
            cloning (LIC) to express 6xHis- and S-tagged proteins with an 
            enterokinase site.
ACCESSION   .
VERSION     .
KEYWORDS    pRSF-2 Ek_LIC
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3609)
  AUTHORS   Novagen (EMD Millipore)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 3609)
  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..3609
                     /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(522..1334)
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
     promoter        complement(1335..1426)
                     /label=AmpR promoter
     rep_origin      complement(1442..2191)
                     /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(2353..2374)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(2390..3469)
                     /label=lacI
                     /note="lac repressor"
     promoter        complement(3470..3547)
                     /label=lacI promoter
     promoter        3593..3609
                     /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 acgacaagct gacgaccggg
      421 tctccgcaag tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
      481 aaatacattc aaatatgtat ccgctcatga attaattctt agaaaaactc atcgagcatc
      541 aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt
      601 ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat
      661 cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa
      721 ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa
      781 agtttatgca tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa
      841 tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg
      901 cggtcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact
      961 gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct
     1021 gttttcccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc
     1081 ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta
     1141 acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc
     1201 ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac
     1261 ccatataaat cagcatccat gttggaattt aatcgcggcc tagagcaaga cgtttcccgt
     1321 tgaatatggc tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     1381 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg catgcagcgc
     1441 tcttccgctt cctcgctcac tgactcgcta cgctcggtcg ttcgactgcg gcgagcggtg
     1501 tcagctcact caaaagcggt aatacggtta tccacagaat caggggataa agccggaaag
     1561 aacatgtgag caaaaagcaa agcaccggaa gaagccaacg ccgcaggcgt ttttccatag
     1621 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agccagaggt ggcgaaaccc
     1681 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1741 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     1801 ttctcatagc tcacgctgtt ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     1861 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     1921 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccattg gtaactgatt
     1981 tagaggactt tgtcttgaag ttatgcacct gttaaggcta aactgaaaga acagattttg
     2041 gtgagtgcgg tcctccaacc cacttacctt ggttcaaaga gttggtagct cagcgaacct
     2101 tgagaaaacc accgttggta gcggtggttt ttctttattt atgagatgat gaatcaatcg
     2161 gtctatcaag tcaacgaaca gctattccgt tactctagat ttcagtgcaa tttatctctt
     2221 caaatgtagc acctgaagtc agccccatac gatataagtt gtaattctca tgttagtcat
     2281 gccccgcgcc caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga
     2341 tcccggtgcc taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt
     2401 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     2461 cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct
     2521 gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc
     2581 ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt
     2641 cggtatcgtc gtatcccact accgagatgt ccgcaccaac gcgcagcccg gactcggtaa
     2701 tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga
     2761 tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt
     2821 cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac
     2881 gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca
     2941 atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt
     3001 tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt
     3061 ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt
     3121 gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg
     3181 acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg
     3241 acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg
     3301 ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt
     3361 tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat
     3421 aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc
     3481 tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga
     3541 tggtgtccgg gatctcgacg ctctccctta tgcgactcct gcattaggaa attaatacga
     3601 ctcactata
//
