LOCUS       40924_19811        4127 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Flexi(R) vector with an ampicillin resistance marker, for mammalian 
            expression from a strongly 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 4127)
  AUTHORS   Promega
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 4127)
  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..4127
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..45
                     /label=CMVd3 promoter
                     /note="truncated human cytomegalovirus (CMV) immediate
                     early promoter that yields strongly reduced expression 
                     relative to the full-length promoter"
     promoter        77..95
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        101..119
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     CDS             158..490
                     /codon_start=1
                     /label=barnase
                     /note="ribonuclease from Bacillus amyloliquefaciens"
                     /translation="MAQVINTFDGVADYLQTYHKLPDNYITKSEAQALGWVASKGNLAD
                     VAPGKSIGGDIFSNREGKLPGKSGRTWREADINYTSGFRNSDRILYSSDWLIYKTTDHY
                     QTFTKIR"
     CDS             533..553
                     /codon_start=1
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
                     /translation="EDLYFQS"
     CDS             566..1453
                     /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(1596..1717)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        1964..2068
                     /label=AmpR promoter
     CDS             2069..2926
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      complement(3087..3675)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    3791..4074
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to
                     maintain plasmids as monomers"
ORIGIN
        1 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt
       61 cagatcacta gaagcttaat acgactcact atagggtacg atttaggtga cactatagaa
      121 taaggagcga tcgccatgga ataagtaagg aatccacatg gcacaggtta tcaacacgtt
      181 tgacggggtt gcggattatc ttcagacata tcataagcta cctgataatt acattacaaa
      241 atcagaagca caagccctcg gctgggtggc atcaaaaggg aaccttgcag acgtcgctcc
      301 ggggaaaagc atcggcggag acatcttctc aaacagggaa ggcaaactcc cgggcaaaag
      361 cggacgaaca tggcgtgaag cggatattaa ctatacatca ggcttcagaa attcagaccg
      421 gattctttac tcaagcgact ggctgattta caaaacaacg gaccattatc agacctttac
      481 aaaaatcaga taatgtttaa tgaccccgtg tcgagctctc gagccaacca ctgaggatct
      541 gtactttcag agcgataacg atggatccga aatcggtact ggctttccat tcgaccccca
      601 ttatgtggaa gtcctgggcg agcgcatgca ctacgtcgat gttggtccgc gcgatggcac
      661 ccctgtgctg ttcctgcacg gtaacccgac ctcctcctac gtgtggcgca acatcatccc
      721 gcatgttgca ccgacccatc gctgcattgc tccagacctg atcggtatgg gcaaatccga
      781 caaaccagac ctgggttatt tcttcgacga ccacgtccgc ttcatggatg ccttcatcga
      841 agccctgggt ctggaagagg tcgtcctggt cattcacgac tggggctccg ctctgggttt
      901 ccactgggcc aagcgcaatc cagagcgcgt caaaggtatt gcatttatgg agttcatccg
      961 ccctatcccg acctgggacg aatggccaga atttgcccgc gagaccttcc aggccttccg
     1021 caccaccgac gtcggccgca agctgatcat cgatcagaac gtttttatcg agggtacgct
     1081 gccgatgggt gtcgtccgcc cgctgactga agtcgagatg gaccattacc gcgagccgtt
     1141 cctgaatcct gttgaccgcg agccactgtg gcgcttccca aacgagctgc caatcgccgg
     1201 tgagccagcg aacatcgtcg cgctggtcga agaatacatg gactggctgc accagtcccc
     1261 tgtcccgaag ctgctgttct ggggcacccc aggcgttctg atcccaccgg ccgaagccgc
     1321 tcgcctggcc aaaagcctgc ctaactgcaa ggctgtggac atcggcccgg gtctgaatct
     1381 gctgcaagaa gacaacccgg acctgatcgg cagcgagatc gcgcgctggc tgtctactct
     1441 ggagatttcc ggttaataga attctagagt cgacctgcag gcatgcaagc tgatccggct
     1501 gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca
     1561 taaccccttg gggcggccgc ttcgagcaga catgataaga tacattgatg agtttggaca
     1621 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc
     1681 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt
     1741 tatgtttcag gttcaggggg agatgtggga ggttttttta agcaagtaaa acctctacaa
     1801 atgtggtaaa atcgaattct aatggatcct ctttgcgctt gcgttttccc ttgtccagat
     1861 agcccagtag ctgacattca tccggggtca gcaccgtttc tgcggactgg ctttctacgt
     1921 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     1981 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     2041 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     2101 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     2161 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     2221 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcactttcaa
     2281 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     2341 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     2401 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     2461 tgcggccaac ttacttctga caactatcgg aggaccgaag gagctaaccg cttttttgca
     2521 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     2581 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     2641 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     2701 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     2761 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     2821 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     2881 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaat tcgaaatgac
     2941 cgaccaagcg acgcccaacc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     3001 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     3061 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     3121 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     3181 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     3241 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     3301 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     3361 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     3421 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     3481 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt
     3541 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     3601 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     3661 aaaaaaggat ttcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     3721 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     3781 cttttatagt ccggaaatac aggaacgcac gctggatggc ccttcgctgg gatggtgaaa
     3841 ccatgaaaaa tggcagcttc agtggattaa gtgggggtaa tgtggcctgt accctctggt
     3901 tgcataggta ttcatacggt taaaatttat caggcgcgat tgcggcagtt tttcgggtgg
     3961 tttgttgcca tttttacctg tctgctgccg tgatcgcgct gaacgcgttt tagcggtgcg
     4021 tacaattaag ggattatggt aaatccactt actgtctgcc ctcgtagcca tcgagataaa
     4081 ccgcagtact ccggccacga tgcgtccggc gtagaggatc gagatct
//
