LOCUS       pCOLA-3-DEST.        5134 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Gateway(R) Nova bacterial destination vector with a ColA origin for 
            expressing proteins tagged at the N-terminus with Strep-Tag II.
ACCESSION   .
VERSION     .
KEYWORDS    pCOLA-3-DEST
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5134)
  AUTHORS   Novagen (EMD Millipore)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 5134)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5134
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(227..1039)
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
     promoter        complement(1040..1131)
                     /label=AmpR promoter
     rep_origin      complement(1149..1784)
                     /direction=LEFT
                     /label=ColA ori
                     /note="Plasmids containing the ColA origin of replication
                     can be propagated in E. coli cells that contain additional 
                     plasmids with compatible origins."
     protein_bind    complement(1948..1969)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(1985..3064)
                     /label=lacI
                     /note="lac repressor"
     promoter        complement(3065..3142)
                     /label=lacI promoter
     promoter        3188..3206
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3207..3231
                     /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             3246..3268
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3275..3277
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     CDS             3284..3307
                     /label=Strep-Tag II
                     /note="peptide that binds Strep-Tactin(R), an engineered 
                     form
                     of streptavidin"
     protein_bind    3317..3441
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        3466..3496
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             3550..4206
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     CDS             4551..4853
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
     protein_bind    complement(4897..5021)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             5023..5040
                     /label=6xHis
                     /note="6xHis affinity tag"
     terminator      5081..5128
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 cctcaggcat ttgagaagca cacggtcaca ctgcttccgg tagtcaataa accggtaaac
       61 cagcaataga cataagcggc tatttaacga ccctgccctg aaccgacgac aagctgacga
      121 ccgggtctcc gcaagtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
      181 tttctaaata cattcaaata tgtatccgct catgaattaa ttcttagaaa aactcatcga
      241 gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat ttttgaaaaa
      301 gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct
      361 ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt
      421 caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg
      481 gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat
      541 caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcgagacgaa
      601 atacgcggtc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac cggcgcagga
      661 acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct aatacctgga
      721 atgctgtttt cccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa
      781 aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg accatctcat
      841 ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg
      901 gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt
      961 tatacccata taaatcagca tccatgttgg aatttaatcg cggcctagag caagacgttt
     1021 cccgttgaat atggctcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     1081 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggcatgc
     1141 tagcgcagaa acgtcctaga agatgccagg aggatactta gcagagagac aataaggccg
     1201 gagcgaagcc gtttttccat aggctccgcc cccctgacga acatcacgaa atctgacgct
     1261 caaatcagtg gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctgatg
     1321 gctccctctt gcgctctcct gttcccgtcc tgcggcgtcc gtgttgtggt ggaggcttta
     1381 cccaaatcac cacgtcccgt tccgtgtaga cagttcgctc caagctgggc tgtgtgcaag
     1441 aaccccccgt tcagcccgac tgctgcgcct tatccggtaa ctatcatctt gagtccaacc
     1501 cggaaagaca cgacaaaacg ccactggcag cagccattgg taactgagaa ttagtggatt
     1561 tagatatcga gagtcttgaa gtggtggcct aacagaggct acactgaaag gacagtattt
     1621 ggtatctgcg ctccactaaa gccagttacc aggttaagca gttccccaac tgacttaacc
     1681 ttcgatcaaa ccgcctcccc aggcggtttt ttcgtttaca gagcaggaga ttacgacgat
     1741 cgtaaaagga tctcaagaag atcctttacg gattcccgac accatcactc tagatttcag
     1801 tgcaatttat ctcttcaaat gtagcacctg aagtcagccc catacgatat aagttgtaat
     1861 tctcatgtta gtcatgcccc gcgcccaccg gaaggagctg actgggttga aggctctcaa
     1921 gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg
     1981 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     2041 acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg
     2101 agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt
     2161 ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat
     2221 aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatgtccgca ccaacgcgca
     2281 gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca
     2341 tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg
     2401 cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat
     2461 gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga
     2521 tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg
     2581 agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat
     2641 tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca
     2701 gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc
     2761 ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg
     2821 ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca
     2881 acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca
     2941 tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc
     3001 gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt
     3061 tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg
     3121 ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga ctcctgcatt
     3181 aggaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc ccctgtagaa
     3241 ataattttgt ttaactttaa taaggagata taccatggca agctggagcc acccgcagtt
     3301 cgaaaagggt gcagtcacaa gtttgtacaa aaaagctgaa cgagaaacgt aaaatgatat
     3361 aaatatcaat atattaaatt agattttgca taaaaaacag actacataat actgtaaaac
     3421 acaacatatc cagtcactat ggcggccgca ttaggcaccc caggctttac actttatgct
     3481 tccggctcgt ataatgtgtg gattttgagt taggatccgt cgagattttc aggagctaag
     3541 gaagctaaaa tggagaaaaa aatcactgga tataccaccg ttgatatatc ccaatggcat
     3601 cgtaaagaac attttgaggc atttcagtca gttgctcaat gtacctataa ccagaccgtt
     3661 cagctggata ttacggcctt tttaaagacc gtaaagaaaa ataagcacaa gttttatccg
     3721 gcctttattc acattcttgc ccgcctgatg aatgctcatc cggaattccg tatggcaatg
     3781 aaagacggtg agctggtgat atgggatagt gttcaccctt gttacaccgt tttccatgag
     3841 caaactgaaa cgttttcatc gctctggagt gaataccacg acgatttccg gcagtttcta
     3901 cacatatatt cgcaagatgt ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg
     3961 tttattgaga atatgttttt cgtctcagcc aatccctggg tgagtttcac cagttttgat
     4021 ttaaacgtgg ccaatatgga caacttcttc gcccccgttt tcactatggg caaatattat
     4081 acgcaaggcg acaaggtgct gatgccgctg gcgattcagg ttcatcatgc cgtttgtgat
     4141 ggcttccatg tcggcagaat gcttaatgaa ttacaacagt actgcgatga gtggcagggc
     4201 ggggcgtaaa cgcgtggatc cggcttacta aaagccagat aacagtatgc gtatttgcgc
     4261 gctgattttt gcggtataag aatatatact gatatgtata cccgaagtat gtcaaaaaga
     4321 ggtatgctat gaagcagcgt attacagtga cagttgacag cgacagctat cagttgctca
     4381 aggcatatat gatgtcaata tctccggtct ggtaagcaca accatgcaga atgaagcccg
     4441 tcgtctgcgt gccgaacgct ggaaagcgga aaatcaggaa gggatggctg aggtcgcccg
     4501 gtttattgaa atgaacggct cttttgctga cgagaacagg ggctggtgaa atgcagttta
     4561 aggtttacac ctataaaaga gagagccgtt atcgtctgtt tgtggatgta cagagtgata
     4621 ttattgacac gcccgggcga cggatggtga tccccctggc cagtgcacgt ctgctgtcag
     4681 ataaagtctc ccgtgaactt tacccggtgg tgcatatcgg ggatgaaagc tggcgcatga
     4741 tgaccaccga tatggccagt gtgccggttt ccgttatcgg ggaagaagtg gctgatctca
     4801 gccaccgcga aaatgacatc aaaaacgcca ttaacctgat gttctgggga atataaatgt
     4861 caggctccct tatacacagc cagtctgcag gtcgaccata gtgactggat atgttgtgtt
     4921 ttacagtatt atgtagtctg ttttttatgc aaaatctaat ttaatatatt gatatttata
     4981 tcattttacg tttctcgttc agctttcttg tacaaagtgg ttcatcacca ccatcatcac
     5041 taagtgatta acctaggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc
     5101 ctctaaacgg gtcttgaggg gttttttgct gaaa
//
