LOCUS       40924_19796        4177 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Flexi(R) vector with a kanamycin resistance marker, for mammalian 
            expression from a slightly 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 4177)
  AUTHORS   Promega
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 4177)
  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..4177
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..100
                     /label=CMVd1 promoter
                     /note="truncated human cytomegalovirus (CMV) immediate
                     early promoter that yields slightly reduced expression 
                     relative to the full-length promoter"
     promoter        132..150
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        156..174
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     CDS             213..545
                     /codon_start=1
                     /label=barnase
                     /note="ribonuclease from Bacillus amyloliquefaciens"
                     /translation="MAQVINTFDGVADYLQTYHKLPDNYITKSEAQALGWVASKGNLAD
                     VAPGKSIGGDIFSNREGKLPGKSGRTWREADINYTSGFRNSDRILYSSDWLIYKTTDHY
                     QTFTKIR"
     CDS             588..608
                     /codon_start=1
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
                     /translation="EDLYFQS"
     CDS             621..1508
                     /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(1651..1772)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             2171..2962
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase from Tn5"
                     /translation="MLEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     rep_origin      complement(3137..3725)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    3841..4124
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to
                     maintain plasmids as monomers"
ORIGIN
        1 ccaaaatcaa cgggactttc caaaatgtcg taataacccc gccccgttga cgcaaatggg
       61 cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat
      121 cactagaagc ttaatacgac tcactatagg gtacgattta ggtgacacta tagaataagg
      181 agcgatcgcc atggaataag taaggaatcc acatggcaca ggttatcaac acgtttgacg
      241 gggttgcgga ttatcttcag acatatcata agctacctga taattacatt acaaaatcag
      301 aagcacaagc cctcggctgg gtggcatcaa aagggaacct tgcagacgtc gctccgggga
      361 aaagcatcgg cggagacatc ttctcaaaca gggaaggcaa actcccgggc aaaagcggac
      421 gaacatggcg tgaagcggat attaactata catcaggctt cagaaattca gaccggattc
      481 tttactcaag cgactggctg atttacaaaa caacggacca ttatcagacc tttacaaaaa
      541 tcagataatg tttaatgacc ccgtgtcgag ctctcgagcc aaccactgag gatctgtact
      601 ttcagagcga taacgatgga tccgaaatcg gtactggctt tccattcgac ccccattatg
      661 tggaagtcct gggcgagcgc atgcactacg tcgatgttgg tccgcgcgat ggcacccctg
      721 tgctgttcct gcacggtaac ccgacctcct cctacgtgtg gcgcaacatc atcccgcatg
      781 ttgcaccgac ccatcgctgc attgctccag acctgatcgg tatgggcaaa tccgacaaac
      841 cagacctggg ttatttcttc gacgaccacg tccgcttcat ggatgccttc atcgaagccc
      901 tgggtctgga agaggtcgtc ctggtcattc acgactgggg ctccgctctg ggtttccact
      961 gggccaagcg caatccagag cgcgtcaaag gtattgcatt tatggagttc atccgcccta
     1021 tcccgacctg ggacgaatgg ccagaatttg cccgcgagac cttccaggcc ttccgcacca
     1081 ccgacgtcgg ccgcaagctg atcatcgatc agaacgtttt tatcgagggt acgctgccga
     1141 tgggtgtcgt ccgcccgctg actgaagtcg agatggacca ttaccgcgag ccgttcctga
     1201 atcctgttga ccgcgagcca ctgtggcgct tcccaaacga gctgccaatc gccggtgagc
     1261 cagcgaacat cgtcgcgctg gtcgaagaat acatggactg gctgcaccag tcccctgtcc
     1321 cgaagctgct gttctggggc accccaggcg ttctgatccc accggccgaa gccgctcgcc
     1381 tggccaaaag cctgcctaac tgcaaggctg tggacatcgg cccgggtctg aatctgctgc
     1441 aagaagacaa cccggacctg atcggcagcg agatcgcgcg ctggctgtct actctggaga
     1501 tttccggtta atagaattct agagtcgacc tgcaggcatg caagctgatc cggctgctaa
     1561 caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc
     1621 ccttggggcg gccgcttcga gcagacatga taagatacat tgatgagttt ggacaaacca
     1681 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat
     1741 ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt
     1801 ttcaggttca gggggagatg tgggaggttt ttttaagcaa gtaaaacctc tacaaatgtg
     1861 gtaaaatcga attctaatgg atcctctttg cgcttgcgtt ttcccttgtc cagatagccc
     1921 agtagctgac attcatccgg ggtcagcacc gtttctgcgg actggctttc tacgtgttcc
     1981 gcttccttta gcagcccttg cgccctgagt gcttgcggca gcgtgagctt caaaagaatt
     2041 gccagctggg gcgccctctg gtaaggttgg gaagccctgc aaagtaaact ggatggcttt
     2101 cttgccgcca aggatctgat ggcgcagggg atcaagatct gatcaagaga caggatgacg
     2161 gtcgtttcgc atgcttgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     2221 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     2281 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     2341 gaatgaactg caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     2401 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     2461 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     2521 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     2581 gaaacatcgc atcgagcgag cacgcactcg gatggaagcc ggtcttgtcg atcaggatga
     2641 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg
     2701 tatgccggat ggtgaggatc tcgtcgtgac tcatggcgat gcctgcttgc cgaatatcat
     2761 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     2821 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     2881 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     2941 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     3001 acgcccaacc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     3061 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     3121 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     3181 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     3241 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     3301 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     3361 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     3421 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     3481 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     3541 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     3601 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     3661 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     3721 ttcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     3781 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttatagt
     3841 ccggaaatac aggaacgcac gctggatggc ccttcgctgg gatggtgaaa ccatgaaaaa
     3901 tggcagcttc agtggattaa gtgggggtaa tgtggcctgt accctctggt tgcataggta
     3961 ttcatacggt taaaatttat caggcgcgat tgcggcagtt tttcgggtgg tttgttgcca
     4021 tttttacctg tctgctgccg tgatcgcgct gaacgcgttt tagcggtgcg tacaattaag
     4081 ggattatggt aaatccactt actgtctgcc ctcgtagcca tcgagataaa ccgcagtact
     4141 ccggccacga tgcgtccggc gtagaggatc gagatct
//
