LOCUS       pEU-His-FV.        4966 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Flexi(R) Vector with the barnase gene, for in vitro transcription 
            and translation of an N-terminally 6xHis-tagged protein.
ACCESSION   .
VERSION     .
KEYWORDS    pEU-His-FV.
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4966)
  AUTHORS   Vinarov DA, Lytle BL, Peterson FC, Tyler EM, Volkman BF, Markley JL.
  TITLE     Cell-free protein production and labeling protocol for NMR-based 
            structural proteomics.
  JOURNAL   Nat. Methods 2004;1:149-53.
  PUBMED    15782178
REFERENCE   2  (bases 1 to 4966)
  AUTHORS   Center for Eukaryotic Structural Genomics
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 4966)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat. 
            Methods"; date: "2004"; volume: "1"; pages: "149-53"
COMMENT     SGRef: number: 2; type: "Journal Article"
COMMENT     To remove the lethal barnase gene, insert an ORF between the SgfI 
            site and the PmeI site or a downstream site.
FEATURES             Location/Qualifiers
     source          1..4966
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        373..391
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     CDS             521..523
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     CDS             527..544
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             576..908
                     /label=barnase
                     /note="ribonuclease from Bacillus amyloliquefaciens"
     promoter        927..957
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             1011..1667
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     misc_feature    1680..1735
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(2425..2441)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2449..2465)
                     /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)."
     promoter        complement(2473..2503)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(2518..2539)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(2827..3415)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3589..4446)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(4447..4551)
                     /label=AmpR promoter
ORIGIN
        1 caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc
       61 atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc
      121 cacgatgcgt ccggcgtaga ggatctggct agcgatgacc ctgctgattg gttcgctgac
      181 catttccggg tgcgggacgg cgttaccaga aactcagaag gttcgtccaa ccaaaccgac
      241 tctggcggca gtttacgaga gagatgatag ggtctgcttc agtaagccag atgctacaca
      301 attaggcttg tacatactgt cgttagaacg cggctacaat taatacataa ccttatgtat
      361 catacacata cgatttaggt gacactatag aatacacgga attcgcgttt ttatttttaa
      421 ttttctttca aatacttcta gctagagtat ttttacaaca attaccaaca acaacaacaa
      481 acaacaacaa cattacattt tacattctac aactacagcc atgggacatc atcatcatca
      541 tcacgcgatc gccatggaat aagtaaagga atcacatggc acaggttatc aacacgtttg
      601 acggggttgc ggattatctt cagacatatc ataagctacc tgataattac attacaaaat
      661 cagaagcaca agccctcggc tgggtggcat caaaagggaa ccttgcagac gtcgctccgg
      721 ggaaaagcat cggcggagac atcttctcaa acagggaagg caaactcccg ggcaaaagcg
      781 gacgaacatg gcgtgaagcg gatattaact atacatcagg cttcagaaat tcagaccgga
      841 ttctttactc aagcgactgg ctgatttaca aaacaacgga ccattatcag acctttacaa
      901 aaatcagata attaggcacc ccaggcttta cactttatgc tttcggctcg tataatgtgt
      961 ggattttgag ttaggatccg tcgagatttt caggagctaa ggaagctaaa atggagaaaa
     1021 aaatcactgg atataccacc gttgatatat cccaatggca tcgtaaagaa cattttgagg
     1081 catttcagtc agttgctcaa tgtacctata accagaccgt tcagctggat attacggcct
     1141 ttttaaagac cgtaaagaaa aataagcaca agttttatcc ggcctttatt cacattcttg
     1201 cccgcctgat gaatgctcat ccggaattcc gtatggcaat gaaagacggt gagctggtga
     1261 tatgggatag tgttcaccct tgttacaccg ttttccatga gcaaactgaa acgttttcat
     1321 cgctctggag tgaataccac gacgatttcc ggcagtttct acacatatat tcgcaagatg
     1381 tggcgtgtta cggtgaaaac ctggcctatt tccctaaagg gtttattgag aatatgtttt
     1441 tcgtctcagc caatccctgg gtgagtttca ccagttttga tttaaacgtg gccaatatgg
     1501 acaacttctt cgcccccgtt ttcaccatgg gcaaatatta tacgcaaggc gacaaggtgc
     1561 tgatgccgct ggcgattcag gttcatcatg ccgtttgtga tggcttccat gtcggcagaa
     1621 tgcttaatga attacaacag tactgcgatg agtggcaggg cggggcgtaa tgtttaaacg
     1681 aattcgagct cggtacccgg ggatcctcta gagtcgacct gcaggcatgc aagctgatcc
     1741 ggctgctaac aaagcccgaa aggaagctga gttggctgct gccaccgctg agcaataact
     1801 agcaaaaaac tagtttacgg ctagcgccgg atgcgacgcc ggtcgcgtct tatccggcct
     1861 tcctatatca ggctgtgttt aagacgccgc cgcttcgccc aaatccctat gccggttcga
     1921 cggctggaca aaatactgtt tatcttccca gcgcaggcag gttaatgtac caccccagca
     1981 gcagccggta tccagcgcgt atataccttc cggcgtacct ttgccctcca gcgatgccca
     2041 gtgaccaaag gcgatgctgt attcttcagc gacagggcca ggaatcgcaa accacggttt
     2101 cagtggggca ggggcctctt ccggcgattc ttactagcta gtatgcatag gtgctgaaat
     2161 ataaagtttg tgtttctaaa acacacgtgg tacgtacgat aacgtacagt gtttttccct
     2221 ccacttaaat cgaagggtag tgtcttggag cgcgcggagt aaacatatat ggttcatata
     2281 tgtccgtagg cacgtaaaaa aagcgaggga ttcgaattcc cccggaaccc ccggttgggg
     2341 cccacgcctc gatcgagcaa aaaaaaaaaa aaagaaaaaa aaaaaaaaaa aaagctttcc
     2401 cgcggccagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     2461 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     2521 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     2581 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     2641 gcgctcttcc gcttcctcac tcactgactc gctgcgctcg gtcgctcggc tgcggcgagc
     2701 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     2761 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     2821 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     2881 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     2941 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     3001 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     3061 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     3121 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     3181 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     3241 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     3301 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     3361 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     3421 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     3481 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     3541 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     3601 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     3661 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     3721 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     3781 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     3841 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     3901 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     3961 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     4021 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     4081 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     4141 ctcaaccaag tcattctgag aatagcgtat gcggcgaccg agttgctctt gcccggcgtc
     4201 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     4261 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     4321 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     4381 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     4441 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     4501 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     4561 ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa
     4621 taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg
     4681 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
     4741 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc
     4801 atcagagcag attgtactga gagtgcacca ttcgacgctc tcccttatgc gactcctgca
     4861 ttaggaagca gcccagtagt aggttgaggc cgttgagcac cgccgccgca aggaatggtg
     4921 catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgc
//
