LOCUS       40924_20306        5064 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Flexi(R) vector with an ampicillin resistance marker, for mammalian 
            expression of a protein with a cleavable N-terminal HaloTag(R).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5064)
  AUTHORS   Promega
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 5064)
  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..5064
                     /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             1067..1957
                     /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             1970..1990
                     /codon_start=1
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
                     /translation="EDLYFQS"
     CDS             2028..2360
                     /codon_start=1
                     /label=barnase
                     /note="ribonuclease from Bacillus amyloliquefaciens"
                     /translation="MAQVINTFDGVADYLQTYHKLPDNYITKSEAQALGWVASKGNLAD
                     VAPGKSIGGDIFSNREGKLPGKSGRTWREADINYTSGFRNSDRILYSSDWLIYKTTDHY
                     QTFTKIR"
     misc_feature    2373..2428
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     polyA_signal    complement(2533..2654)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        2901..3005
                     /label=AmpR promoter
     CDS             3006..3863
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      complement(4024..4612)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    4728..5011
                     /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 ggctagcaaa gccaccatgg cagaaatcgg
     1081 tactggcttt ccattcgacc cccattatgt ggaagtcctg ggcgagcgca tgcactacgt
     1141 cgatgttggt ccgcgcgatg gcacccctgt gctgttcctg cacggtaacc cgacctcctc
     1201 ctacgtgtgg cgcaacatca tcccgcatgt tgcaccgacc catcgctgca ttgctccaga
     1261 cctgatcggt atgggcaaat ccgacaaacc agacctgggt tatttcttcg acgaccacgt
     1321 ccgcttcatg gatgccttca tcgaagccct gggtctggaa gaggtcgtcc tggtcattca
     1381 cgactggggc tccgctctgg gtttccactg ggccaagcgc aatccagagc gcgtcaaagg
     1441 tattgcattt atggagttca tccgccctat cccgacctgg gacgaatggc cagaatttgc
     1501 ccgcgagacc ttccaggcct tccgcaccac cgacgtcggc cgcaagctga tcatcgatca
     1561 gaacgttttt atcgagggta cgctgccgat gggtgtcgtc cgcccgctga ctgaagtcga
     1621 gatggaccat taccgcgagc cgttcctgaa tcctgttgac cgcgagccac tgtggcgctt
     1681 cccaaacgag ctgccaatcg ccggtgagcc agcgaacatc gtcgcgctgg tcgaagaata
     1741 catggactgg ctgcaccagt cccctgtccc gaagctgctg ttctggggca ccccaggcgt
     1801 tctgatccca ccggccgaag ccgctcgcct ggccaaaagc ctgcctaact gcaaggctgt
     1861 ggacatcggc ccgggtctga atctgctgca agaagacaac ccggacctga tcggcagcga
     1921 gatcgcgcgc tggctgtcga cgctcgagat ttccggcgag ccaaccactg aggatctgta
     1981 ctttcagagc gataacgcga tcgccatgga ataagtaagg aatccacatg gcacaggtta
     2041 tcaacacgtt tgacggggtt gcggattatc ttcagacata tcataagcta cctgataatt
     2101 acattacaaa atcagaagca caagccctcg gctgggtggc atcaaaaggg aaccttgcag
     2161 acgtcgctcc ggggaaaagc atcggcggag acatcttctc aaacagggaa ggcaaactcc
     2221 cgggcaaaag cggacgaaca tggcgtgaag cggatattaa ctatacatca ggcttcagaa
     2281 attcagaccg gattctttac tcaagcgact ggctgattta caaaacaacg gaccattatc
     2341 agacctttac aaaaatcaga taatgtttaa acgaattcgg gctcggtacc cggggatcct
     2401 ctagagtcga cctgcaggca tgcaagctga tccggctgct aacaaagccc gaaaggaagc
     2461 tgagttggct gctgccaccg ctgagcaata actagcataa ccccttgggg cggccgcttc
     2521 gagcagacat gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa
     2581 aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     2641 gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggaga
     2701 tgtgggaggt ttttttaagc aagtaaaacc tctacaaatg tggtaaaatc gaattctaat
     2761 ggatcctctt tgcgcttgcg ttttcccttg tccagatagc ccagtagctg acattcatcc
     2821 ggggtcagca ccgtttctgc ggactggctt tctacgtaat ggtttcttag acgtcaggtg
     2881 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     2941 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     3001 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     3061 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     3121 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     3181 gccccgaaga acgttttcca atgatgagca ctttcaaagt tctgctatgt ggcgcggtat
     3241 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     3301 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     3361 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     3421 ctatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     3481 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     3541 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     3601 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     3661 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     3721 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     3781 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     3841 gtgcctcact gattaagcat tggtaattcg aaatgaccga ccaagcgacg cccaaccggt
     3901 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     3961 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     4021 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4081 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4141 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     4201 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4261 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     4321 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     4381 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     4441 gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt
     4501 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     4561 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatttc aagaagatcc
     4621 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     4681 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttatagtccg gaaatacagg
     4741 aacgcacgct ggatggccct tcgctgggat ggtgaaacca tgaaaaatgg cagcttcagt
     4801 ggattaagtg ggggtaatgt ggcctgtacc ctctggttgc ataggtattc atacggttaa
     4861 aatttatcag gcgcgattgc ggcagttttt cgggtggttt gttgccattt ttacctgtct
     4921 gctgccgtga tcgcgctgaa cgcgttttag cggtgcgtac aattaaggga ttatggtaaa
     4981 tccacttact gtctgccctc gtagccatcg agataaaccg cagtactccg gccacgatgc
     5041 gtccggcgta gaggatcgag atct
//
