LOCUS       40924_19801        4134 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Flexi(R) vector with an ampicillin resistance marker, for mammalian 
            expression from a moderately compromised CMV promoter of a protein 
            with a cleavable C-terminal HaloTag(R).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4134)
  AUTHORS   Promega
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 4134)
  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..4134
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..52
                     /label=CMVd2 promoter
                     /note="truncated human cytomegalovirus (CMV) immediate
                     early promoter that yields moderately reduced expression 
                     relative to the full-length promoter"
     promoter        84..102
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        108..126
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     CDS             165..497
                     /codon_start=1
                     /label=barnase
                     /note="ribonuclease from Bacillus amyloliquefaciens"
                     /translation="MAQVINTFDGVADYLQTYHKLPDNYITKSEAQALGWVASKGNLAD
                     VAPGKSIGGDIFSNREGKLPGKSGRTWREADINYTSGFRNSDRILYSSDWLIYKTTDHY
                     QTFTKIR"
     CDS             540..560
                     /codon_start=1
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
                     /translation="EDLYFQS"
     CDS             573..1460
                     /codon_start=1
                     /label=HaloTag(R)
                     /note="modified bacterial dehalogenase that forms covalent
                     bonds with chloroalkane derivatives"
                     /translation="SEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSY
                     VWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIH
                     DWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIID
                     QNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVE
                     EYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLI
                     GSEIARWLSTLEISG"
     polyA_signal    complement(1603..1724)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        1971..2075
                     /label=AmpR promoter
     CDS             2076..2933
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      complement(3094..3682)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    3798..4081
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to
                     maintain plasmids as monomers"
ORIGIN
        1 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt
       61 gaaccgtcag atcactagaa gcttaatacg actcactata gggtacgatt taggtgacac
      121 tatagaataa ggagcgatcg ccatggaata agtaaggaat ccacatggca caggttatca
      181 acacgtttga cggggttgcg gattatcttc agacatatca taagctacct gataattaca
      241 ttacaaaatc agaagcacaa gccctcggct gggtggcatc aaaagggaac cttgcagacg
      301 tcgctccggg gaaaagcatc ggcggagaca tcttctcaaa cagggaaggc aaactcccgg
      361 gcaaaagcgg acgaacatgg cgtgaagcgg atattaacta tacatcaggc ttcagaaatt
      421 cagaccggat tctttactca agcgactggc tgatttacaa aacaacggac cattatcaga
      481 cctttacaaa aatcagataa tgtttaatga ccccgtgtcg agctctcgag ccaaccactg
      541 aggatctgta ctttcagagc gataacgatg gatccgaaat cggtactggc tttccattcg
      601 acccccatta tgtggaagtc ctgggcgagc gcatgcacta cgtcgatgtt ggtccgcgcg
      661 atggcacccc tgtgctgttc ctgcacggta acccgacctc ctcctacgtg tggcgcaaca
      721 tcatcccgca tgttgcaccg acccatcgct gcattgctcc agacctgatc ggtatgggca
      781 aatccgacaa accagacctg ggttatttct tcgacgacca cgtccgcttc atggatgcct
      841 tcatcgaagc cctgggtctg gaagaggtcg tcctggtcat tcacgactgg ggctccgctc
      901 tgggtttcca ctgggccaag cgcaatccag agcgcgtcaa aggtattgca tttatggagt
      961 tcatccgccc tatcccgacc tgggacgaat ggccagaatt tgcccgcgag accttccagg
     1021 ccttccgcac caccgacgtc ggccgcaagc tgatcatcga tcagaacgtt tttatcgagg
     1081 gtacgctgcc gatgggtgtc gtccgcccgc tgactgaagt cgagatggac cattaccgcg
     1141 agccgttcct gaatcctgtt gaccgcgagc cactgtggcg cttcccaaac gagctgccaa
     1201 tcgccggtga gccagcgaac atcgtcgcgc tggtcgaaga atacatggac tggctgcacc
     1261 agtcccctgt cccgaagctg ctgttctggg gcaccccagg cgttctgatc ccaccggccg
     1321 aagccgctcg cctggccaaa agcctgccta actgcaaggc tgtggacatc ggcccgggtc
     1381 tgaatctgct gcaagaagac aacccggacc tgatcggcag cgagatcgcg cgctggctgt
     1441 ctactctgga gatttccggt taatagaatt ctagagtcga cctgcaggca tgcaagctga
     1501 tccggctgct aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata
     1561 actagcataa ccccttgggg cggccgcttc gagcagacat gataagatac attgatgagt
     1621 ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg
     1681 ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca
     1741 ttcattttat gtttcaggtt cagggggaga tgtgggaggt ttttttaagc aagtaaaacc
     1801 tctacaaatg tggtaaaatc gaattctaat ggatcctctt tgcgcttgcg ttttcccttg
     1861 tccagatagc ccagtagctg acattcatcc ggggtcagca ccgtttctgc ggactggctt
     1921 tctacgtaat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     1981 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     2041 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     2101 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     2161 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     2221 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     2281 ctttcaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     2341 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     2401 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     2461 ataacactgc ggccaactta cttctgacaa ctatcggagg accgaaggag ctaaccgctt
     2521 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     2581 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     2641 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     2701 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     2761 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     2821 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     2881 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaattcg
     2941 aaatgaccga ccaagcgacg cccaaccggt atcagctcac tcaaaggcgg taatacggtt
     3001 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     3061 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     3121 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     3181 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     3241 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     3301 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     3361 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     3421 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     3481 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     3541 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3601 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3661 gcgcagaaaa aaaggatttc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     3721 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     3781 ctagatcctt ttatagtccg gaaatacagg aacgcacgct ggatggccct tcgctgggat
     3841 ggtgaaacca tgaaaaatgg cagcttcagt ggattaagtg ggggtaatgt ggcctgtacc
     3901 ctctggttgc ataggtattc atacggttaa aatttatcag gcgcgattgc ggcagttttt
     3961 cgggtggttt gttgccattt ttacctgtct gctgccgtga tcgcgctgaa cgcgttttag
     4021 cggtgcgtac aattaaggga ttatggtaaa tccacttact gtctgccctc gtagccatcg
     4081 agataaaccg cagtactccg gccacgatgc gtccggcgta gaggatcgag atct
//
