LOCUS       40924_19841        4312 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Flexi(R) vector with an ampicillin resistance marker, for bacterial 
            or cell-free expression of a protein with a cleavable C-terminal 
            HaloTag(R)-6xHis dual tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4312)
  AUTHORS   Promega
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 4312)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Subclone a coding sequence between the SgfI/AsiSI and 
            EcoICRI/Eco53kI sites to remove the lethal barnase gene.
FEATURES             Location/Qualifiers
     source          1..4312
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..39
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             92..424
                     /codon_start=1
                     /label=barnase
                     /note="ribonuclease from Bacillus amyloliquefaciens"
                     /translation="MAQVINTFDGVADYLQTYHKLPDNYITKSEAQALGWVASKGNLAD
                     VAPGKSIGGDIFSNREGKLPGKSGRTWREADINYTSGFRNSDRILYSSDWLIYKTTDHY
                     QTFTKIR"
     CDS             467..487
                     /codon_start=1
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
                     /translation="EDLYFQS"
     CDS             500..1387
                     /codon_start=1
                     /label=HaloTag(R)
                     /note="modified bacterial dehalogenase that forms covalent
                     bonds with chloroalkane derivatives"
                     /translation="SEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSY
                     VWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIH
                     DWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIID
                     QNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVE
                     EYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLI
                     GSEIARWLSTLEISG"
     CDS             1388..1405
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     terminator      1430..1477
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        1706..1810
                     /label=AmpR promoter
     CDS             1811..2668
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      complement(2829..3417)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    3533..3816
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to
                     maintain plasmids as monomers"
     terminator      4046..4132
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      4224..4251
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
ORIGIN
        1 ggatctcgat cccgcgaaat taatacgact cactataggg tattttaact ttactaagga
       61 gcgatcgcca tggaataagt aaggaatcca catggcacag gttatcaaca cgtttgacgg
      121 ggttgcggat tatcttcaga catatcataa gctacctgat aattacatta caaaatcaga
      181 agcacaagcc ctcggctggg tggcatcaaa agggaacctt gcagacgtcg ctccggggaa
      241 aagcatcggc ggagacatct tctcaaacag ggaaggcaaa ctcccgggca aaagcggacg
      301 aacatggcgt gaagcggata ttaactatac atcaggcttc agaaattcag accggattct
      361 ttactcaagc gactggctga tttacaaaac aacggaccat tatcagacct ttacaaaaat
      421 cagataatgt ttaatgaccc cgtgtcgagc tctcgagcca accactgagg atctgtactt
      481 tcagagcgat aacgatggat ccgaaatcgg tactggcttt ccattcgacc cccattatgt
      541 ggaagtcctg ggcgagcgca tgcactacgt cgatgttggt ccgcgcgatg gcacccctgt
      601 gctgttcctg cacggtaacc cgacctcctc ctacgtgtgg cgcaacatca tcccgcatgt
      661 tgcaccgacc catcgctgca ttgctccaga cctgatcggt atgggcaaat ccgacaaacc
      721 agacctgggt tatttcttcg acgaccacgt ccgcttcatg gatgccttca tcgaagccct
      781 gggtctggaa gaggtcgtcc tggtcattca cgactggggc tccgctctgg gtttccactg
      841 ggccaagcgc aatccagagc gcgtcaaagg tattgcattt atggagttca tccgccctat
      901 cccgacctgg gacgaatggc cagaatttgc ccgcgagacc ttccaggcct tccgcaccac
      961 cgacgtcggc cgcaagctga tcatcgatca gaacgttttt atcgagggta cgctgccgat
     1021 gggtgtcgtc cgcccgctga ctgaagtcga gatggaccat taccgcgagc cgttcctgaa
     1081 tcctgttgac cgcgagccac tgtggcgctt cccaaacgag ctgccaatcg ccggtgagcc
     1141 agcgaacatc gtcgcgctgg tcgaagaata catggactgg ctgcaccagt cccctgtccc
     1201 gaagctgctg ttctggggca ccccaggcgt tctgatccca ccggccgaag ccgctcgcct
     1261 ggccaaaagc ctgcctaact gcaaggctgt ggacatcggc ccgggtctga atctgctgca
     1321 agaagacaac ccggacctga tcggcagcga gatcgcgcgc tggctgtcta ctctggagat
     1381 ttccggtcat catcaccatc accactaata gtcaaccgct gagcaataac tagcataacc
     1441 ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg
     1501 ttcgcttgct gtccataaaa ccgcccagtc tagctatcgc catgtaagcc cactgcaagc
     1561 tacctgcttt ctctttgcgc ttgcgttttc ccttgtccag atagcccagt agctgacatt
     1621 catccggggt cagcaccgtt tctgcggact ggctttctac gtaatggttt cttagacgtc
     1681 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     1741 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     1801 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     1861 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     1921 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     1981 ttttcgcccc gaagaacgtt ttccaatgat gagcactttc aaagttctgc tatgtggcgc
     2041 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     2101 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     2161 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     2221 gacaactatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     2281 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     2341 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     2401 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     2461 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     2521 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     2581 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     2641 gataggtgcc tcactgatta agcattggta attcgaaatg accgaccaag cgacgcccaa
     2701 ccggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     2761 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     2821 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     2881 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     2941 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     3001 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     3061 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     3121 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     3181 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     3241 tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag
     3301 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     3361 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atttcaagaa
     3421 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     3481 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttata gtccggaaat
     3541 acaggaacgc acgctggatg gcccttcgct gggatggtga aaccatgaaa aatggcagct
     3601 tcagtggatt aagtgggggt aatgtggcct gtaccctctg gttgcatagg tattcatacg
     3661 gttaaaattt atcaggcgcg attgcggcag tttttcgggt ggtttgttgc catttttacc
     3721 tgtctgctgc cgtgatcgcg ctgaacgcgt tttagcggtg cgtacaatta agggattatg
     3781 gtaaatccac ttactgtctg ccctcgtagc catcgagata aaccgcagta ctccggccac
     3841 gatgcgtccg gcgtagagga tcgagatctt ttcagcctga tacagattaa atcagaacgc
     3901 agaagcggtc tgataaaaca gaatttgcct ggcggcagta gcgcggtggt cccacctgac
     3961 cccatgccga actcagaagt gaaacgccgt agcgccgatg gtagtgtggg gtctccccat
     4021 gcgagagtag ggaactgcca ggcatcaaat aaaacgaaag gctcagtcga aagactgggc
     4081 ctttcgtttt atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa atccgccggg
     4141 agcggatttg aacgttgcga agcaacggcc cggagggtgg cgggcaggac gcccgccata
     4201 aactgccagg catcaaatta agcagaaggc catcctgacg gatggccttt ttgcgtttct
     4261 acaaactctt ttgtttattt ttctaaatac attcaaatat gtatccgctc at
//
