LOCUS       40924_20321        4183 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 an N-terminal HaloTag(R).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4183)
  AUTHORS   Promega
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 4183)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Subclone a coding sequence between the SgfI/AsiSI and PmeI sites to 
            remove the lethal barnase gene.
FEATURES             Location/Qualifiers
     source          1..4183
                     /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             191..1081
                     /codon_start=1
                     /label=HaloTag(R)
                     /note="modified bacterial dehalogenase that forms covalent
                     bonds with chloroalkane derivatives"
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             1094..1114
                     /codon_start=1
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
                     /translation="EDLYFQS"
     CDS             1152..1484
                     /codon_start=1
                     /label=barnase
                     /note="ribonuclease from Bacillus amyloliquefaciens"
                     /translation="MAQVINTFDGVADYLQTYHKLPDNYITKSEAQALGWVASKGNLAD
                     VAPGKSIGGDIFSNREGKLPGKSGRTWREADINYTSGFRNSDRILYSSDWLIYKTTDHY
                     QTFTKIR"
     misc_feature    1497..1552
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     polyA_signal    complement(1657..1778)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             2177..2968
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase from Tn5"
                     /translation="MLEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     rep_origin      complement(3143..3731)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    3847..4130
                     /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 agaattcatc atggcagaaa tcggtactgg ctttccattc gacccccatt atgtggaagt
      241 cctgggcgag cgcatgcact acgtcgatgt tggtccgcgc gatggcaccc ctgtgctgtt
      301 cctgcacggt aacccgacct cctcctacgt gtggcgcaac atcatcccgc atgttgcacc
      361 gacccatcgc tgcattgctc cagacctgat cggtatgggc aaatccgaca aaccagacct
      421 gggttatttc ttcgacgacc acgtccgctt catggatgcc ttcatcgaag ccctgggtct
      481 ggaagaggtc gtcctggtca ttcacgactg gggctccgct ctgggtttcc actgggccaa
      541 gcgcaatcca gagcgcgtca aaggtattgc atttatggag ttcatccgcc ctatcccgac
      601 ctgggacgaa tggccagaat ttgcccgcga gaccttccag gccttccgca ccaccgacgt
      661 cggccgcaag ctgatcatcg atcagaacgt ttttatcgag ggtacgctgc cgatgggtgt
      721 cgtccgcccg ctgactgaag tcgagatgga ccattaccgc gagccgttcc tgaatcctgt
      781 tgaccgcgag ccactgtggc gcttcccaaa cgagctgcca atcgccggtg agccagcgaa
      841 catcgtcgcg ctggtcgaag aatacatgga ctggctgcac cagtcccctg tcccgaagct
      901 gctgttctgg ggcaccccag gcgttctgat cccaccggcc gaagccgctc gcctggccaa
      961 aagcctgcct aactgcaagg ctgtggacat cggcccgggt ctgaatctgc tgcaagaaga
     1021 caacccggac ctgatcggca gcgagatcgc gcgctggctg tcgacgctcg agatttccgg
     1081 cgagccaacc actgaggatc tgtactttca gagcgataac gcgatcgcca tggaataagt
     1141 aaggaatcca catggcacag gttatcaaca cgtttgacgg ggttgcggat tatcttcaga
     1201 catatcataa gctacctgat aattacatta caaaatcaga agcacaagcc ctcggctggg
     1261 tggcatcaaa agggaacctt gcagacgtcg ctccggggaa aagcatcggc ggagacatct
     1321 tctcaaacag ggaaggcaaa ctcccgggca aaagcggacg aacatggcgt gaagcggata
     1381 ttaactatac atcaggcttc agaaattcag accggattct ttactcaagc gactggctga
     1441 tttacaaaac aacggaccat tatcagacct ttacaaaaat cagataatgt ttaaacgaat
     1501 tcgggctcgg tacccgggga tcctctagag tcgacctgca ggcatgcaag ctgatccggc
     1561 tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc accgctgagc aataactagc
     1621 ataacccctt ggggcggccg cttcgagcag acatgataag atacattgat gagtttggac
     1681 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     1741 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
     1801 ttatgtttca ggttcagggg gagatgtggg aggttttttt aagcaagtaa aacctctaca
     1861 aatgtggtaa aatcgaattc taatggatcc tctttgcgct tgcgttttcc cttgtccaga
     1921 tagcccagta gctgacattc atccggggtc agcaccgttt ctgcggactg gctttctacg
     1981 tgttccgctt cctttagcag cccttgcgcc ctgagtgctt gcggcagcgt gagcttcaaa
     2041 agaattgcca gctggggcgc cctctggtaa ggttgggaag ccctgcaaag taaactggat
     2101 ggctttcttg ccgccaagga tctgatggcg caggggatca agatctgatc aagagacagg
     2161 atgacggtcg tttcgcatgc ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     2221 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     2281 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     2341 tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     2401 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     2461 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     2521 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     2581 ccaagcgaaa catcgcatcg agcgagcacg cactcggatg gaagccggtc ttgtcgatca
     2641 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     2701 ggcgcgtatg ccggatggtg aggatctcgt cgtgactcat ggcgatgcct gcttgccgaa
     2761 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     2821 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     2881 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     2941 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     3001 caagcgacgc ccaaccggta tcagctcact caaaggcggt aatacggtta tccacagaat
     3061 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     3121 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     3181 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     3241 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     3301 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     3361 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     3421 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     3481 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     3541 cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct
     3601 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     3661 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     3721 aaggatttca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     3781 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     3841 tatagtccgg aaatacagga acgcacgctg gatggccctt cgctgggatg gtgaaaccat
     3901 gaaaaatggc agcttcagtg gattaagtgg gggtaatgtg gcctgtaccc tctggttgca
     3961 taggtattca tacggttaaa atttatcagg cgcgattgcg gcagtttttc gggtggtttg
     4021 ttgccatttt tacctgtctg ctgccgtgat cgcgctgaac gcgttttagc ggtgcgtaca
     4081 attaagggat tatggtaaat ccacttactg tctgccctcg tagccatcga gataaaccgc
     4141 agtactccgg ccacgatgcg tccggcgtag aggatcgaga tct
//
