LOCUS       40924_19836        5527 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Flexi(R) vector with a kanamycin/G418 resistance marker, for 
            mammalian expression 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 5527)
  AUTHORS   Promega
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 5527)
  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..5527
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        139..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     promoter        1033..1051
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1087..1419
                     /codon_start=1
                     /label=barnase
                     /note="ribonuclease from Bacillus amyloliquefaciens"
                     /translation="MAQVINTFDGVADYLQTYHKLPDNYITKSEAQALGWVASKGNLAD
                     VAPGKSIGGDIFSNREGKLPGKSGRTWREADINYTSGFRNSDRILYSSDWLIYKTTDHY
                     QTFTKIR"
     CDS             1462..1482
                     /codon_start=1
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
                     /translation="EDLYFQS"
     CDS             1495..2382
                     /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(2525..2646)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        2891..3248
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     promoter        3263..3310
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter"
     CDS             3342..4133
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    4200..4248
                     /label=poly(A) signal
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(4487..5075)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    5191..5474
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to
                     maintain plasmids as monomers"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tggtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt attaatacga ctcactatag ggctagcgat cgccatggaa taagtaagga
     1081 atccacatgg cacaggttat caacacgttt gacggggttg cggattatct tcagacatat
     1141 cataagctac ctgataatta cattacaaaa tcagaagcac aagccctcgg ctgggtggca
     1201 tcaaaaggga accttgcaga cgtcgctccg gggaaaagca tcggcggaga catcttctca
     1261 aacagggaag gcaaactccc gggcaaaagc ggacgaacat ggcgtgaagc ggatattaac
     1321 tatacatcag gcttcagaaa ttcagaccgg attctttact caagcgactg gctgatttac
     1381 aaaacaacgg accattatca gacctttaca aaaatcagat aatgtttaat gaccccgtgt
     1441 cgagctctcg agccaaccac tgaggatctg tactttcaga gcgataacga tggatccgaa
     1501 atcggtactg gctttccatt cgacccccat tatgtggaag tcctgggcga gcgcatgcac
     1561 tacgtcgatg ttggtccgcg cgatggcacc cctgtgctgt tcctgcacgg taacccgacc
     1621 tcctcctacg tgtggcgcaa catcatcccg catgttgcac cgacccatcg ctgcattgct
     1681 ccagacctga tcggtatggg caaatccgac aaaccagacc tgggttattt cttcgacgac
     1741 cacgtccgct tcatggatgc cttcatcgaa gccctgggtc tggaagaggt cgtcctggtc
     1801 attcacgact ggggctccgc tctgggtttc cactgggcca agcgcaatcc agagcgcgtc
     1861 aaaggtattg catttatgga gttcatccgc cctatcccga cctgggacga atggccagaa
     1921 tttgcccgcg agaccttcca ggccttccgc accaccgacg tcggccgcaa gctgatcatc
     1981 gatcagaacg tttttatcga gggtacgctg ccgatgggtg tcgtccgccc gctgactgaa
     2041 gtcgagatgg accattaccg cgagccgttc ctgaatcctg ttgaccgcga gccactgtgg
     2101 cgcttcccaa acgagctgcc aatcgccggt gagccagcga acatcgtcgc gctggtcgaa
     2161 gaatacatgg actggctgca ccagtcccct gtcccgaagc tgctgttctg gggcacccca
     2221 ggcgttctga tcccaccggc cgaagccgct cgcctggcca aaagcctgcc taactgcaag
     2281 gctgtggaca tcggcccggg tctgaatctg ctgcaagaag acaacccgga cctgatcggc
     2341 agcgagatcg cgcgctggct gtctactctg gagatttccg gttaatagaa ttctagagtc
     2401 gacctgcagg catgcaagct gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
     2461 ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcggccgct tcgagcagac
     2521 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
     2581 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
     2641 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag
     2701 gtttttttaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgaatttta acaaaatatt
     2761 aacgcttaca atttcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     2821 cgcatacgcg gatctgcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt
     2881 taggtacctt ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa
     2941 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3001 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3061 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     3121 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     3181 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     3241 tttgcaaaaa gcttaattaa ctgttgacaa ttaatcatcg gcatagtata tcggcatagt
     3301 ataatacgac aaggtgagga actaaaccca ggaggcagat catgattgaa caagatggat
     3361 tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac
     3421 agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc
     3481 tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc
     3541 tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag
     3601 cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc
     3661 ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg
     3721 atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc
     3781 ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc
     3841 cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga
     3901 cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca
     3961 tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg
     4021 atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg
     4081 ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg
     4141 gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gatggccgca
     4201 ataaaatatc tttattttca ttacatctgt gtgttggttt tttgtgtgaa tcgatagcga
     4261 taaggatcct ctttgcgctt gcgttttccc ttgtccagat agcccagtag ctgacattca
     4321 tccggggtca gcaccgtttc tgcggactgg ctttctaccc ggtatcagct cactcaaagg
     4381 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     4441 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     4501 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     4561 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     4621 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     4681 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     4741 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     4801 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4861 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4921 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4981 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     5041 agcagcagat tacgcgcaga aaaaaaggat ttcaagaaga tcctttgatc ttttctacgg
     5101 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     5161 aaaggatctt cacctagatc cttttatagt ccggaaatac aggaacgcac gctggatggc
     5221 ccttcgctgg gatggtgaaa ccatgaaaaa tggcagcttc agtggattaa gtgggggtaa
     5281 tgtggcctgt accctctggt tgcataggta ttcatacggt taaaatttat caggcgcgat
     5341 tgcggcagtt tttcgggtgg tttgttgcca tttttacctg tctgctgccg tgatcgcgct
     5401 gaacgcgttt tagcggtgcg tacaattaag ggattatggt aaatccactt actgtctgcc
     5461 ctcgtagcca tcgagataaa ccgcagtact ccggccacga tgcgtccggc gtagaggatc
     5521 gagatct
//
