LOCUS       V032227                 9613 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V032227
VERSION     V032227
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     enhancer        235..614
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        615..818
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     promoter        863..881
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /label="T7 promoter"
     CDS             964..5103
                     /label="Map3k5(NM_008580)"
                     /note="Map3k5(NM_008580)"
                     /gene="Map3k5"
     CDS             5110..5175
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     polyA_signal    5213..5437
                     /note="bovine growth hormone polyadenylation signal"
                     /label="bGH poly(A) signal"
     rep_origin      5483..5911
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        5925..6254
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             6321..7115
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /note="confers resistance to neomycin, kanamycin, and G418
                     (Geneticin®)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /transl_table=1
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*"
                     /label="NeoR/KanR"
     polyA_signal    7289..7410
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(7459..7475)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    7483..7499
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be
                     relieved by adding lactose or
                     isopropyl-β-D-thiogalactopyranoside (IPTG)."
                     /label="lac operator"
     promoter        complement(7507..7537)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    7552..7573
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7861..8446)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8617..9477)
                     /codon_start=1
                     /gene="bla"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /product="β-lactamase"
                     /transl_table=1
                     /translation="MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEY
                     SPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDR
                     WEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRS
                     ALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGA
                     SLIKHW*"
                     /label="AmpR"
     promoter        complement(9478..9582)
                     /gene="bla"
                     /label="AmpR promoter"
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattcgcc
      961 accatgggca ccgaagccgg cgagggcatc actttctccg tgccgccctt tgcgtccgtg
     1021 ggcttttgca ccatcccgga gggcggcagc tgccggaggg gaggaggagc ggcgacggcg
     1081 gcggagggag agccgagcct gcagcccttg cttgtgccgc cgccgccgcc accgccgggc
     1141 agtttctgga acgtggagag cgcggcggct cccggcacca gctgccctac caccgccccc
     1201 gggagcagcg ccacccgagg ccggggcaac tctggcagcg ggggcggccg gcgtaccacc
     1261 gtggcttatg tgatcaatga ggcgagccag gggcagctgg tggtggcgga gagcgaggcc
     1321 ctgcagagcc tgcgggaggc gtgcgaggcg gtgggcgcca ccctggagac cctgcatttc
     1381 gggaagctgg acttcgggga aaccgccgtg ctggaccgtt tttacaacgc agatattgct
     1441 gtggtggaga tgagcgacgc cttccggcag ccttccttgt tttaccatct tggagtgcga
     1501 gaaagcttca gtatggccaa caacatcatc ctctactgcg atactaattc cgattcactc
     1561 cagtccctga aggaaattat ttgccagaag aatactgtgt gcaccgggaa ctacaccttc
     1621 atcccctaca tggtgacacc acacaacaag gtctactgct gtgacagcag cttcatgaag
     1681 ggcctgactg agctcatgca gcctaacttc gagctcctgc tggggcccat ctgcttaccc
     1741 ctcgtggacc gtttcgttca gcttttgaag gtggcgcaag cgagttccag ccagtacttc
     1801 cgggaatcca tactcagcga tatcaggaaa gcacggaacc tgtacactgg taaagaattg
     1861 gcagctgaat tggcgaggat ccggcaacgg gtagacaata ttgaagtttt gacagcagac
     1921 attgttatca acctgttgct ttcctacaga gacatccagg actatgactc catagtgaag
     1981 ctggtagaga cgttagaaaa acttccaacc ttcgacttgg cctcgcacca ccacgtgaag
     2041 tttcattacg catttgcact gaacaggaga aatctccctg gagacagagc aaaagctctt
     2101 gacatcatga tccccatggt acaaagtgaa gagcaagttg cttcggatat gtattgtctc
     2161 gttggccgaa tctacaaaga tatgtttttg gattcaaatt tcactgacac ggaaagcagg
     2221 gaccatggag catcttggtt caaaaaggca tttgagtctg agccaacact acagtcagga
     2281 attaattatg cagttctctt gctggcggct gggcaccagt ttgaatcttc ttttgagctt
     2341 cggaaagttg gagtgaagct gagtagcctt ctgggtaaaa aggggaactt ggaaaaactc
     2401 cagagctact gggaagtcgg attttttctg ggagccagtg tcctggccaa cgaccacctc
     2461 agggtcattc aggcatccga gaagctgttc agactgaaga caccagcgtg gtacctcaag
     2521 tctatcgtgg agaccattct gatatataaa cactttgtga aactgaccac agagcagccc
     2581 tcggctaagc aggaactcgt ggacttctgg atggatttcc tggtcgaggc cacaaaaaca
     2641 gatgttactg ttgtcaggtt cccagtatta atattagagc caaccaaaat ctatcagcct
     2701 tcttacctgt ctatcaataa tgaagttgag gagaagacaa tatctatttg gcatgttctc
     2761 cctgatgaca agaaaggtat acacgagtgg aacttcggtg cttcctctgt caggggagtg
     2821 agcatctcta agtttgaaga acgatgctgt ttcctttatg tgcttcacaa ttctgatgac
     2881 ttccaaatct acttctgcac agaacttcac tgtaaaaggt tttttgaaat ggtgaatacc
     2941 atcacggaag agaaggggag aggcgcggag gacggagact gtgagggtga ctccctggag
     3001 tatgactacg aatacgatga aaatggagac agagttgtgt tagggaaggg cacttatggg
     3061 atcgtctatg caggacgaga cctgagcaac caggtcagaa ttgctattaa ggaaatccca
     3121 gagagagaca gcagatactc tcagcccctg catgaagaaa tcgccctgca caagcatctg
     3181 aaacacaaaa acattgtcca gtatctgggc tccttcagtg agaacggctt catcaaaatc
     3241 ttcatggagc aagtcccagg aggaagcctc tctgctctcc ttcgttcaaa atggggccca
     3301 ttaaaggaca acgaacagac gattggcttc tatacgaagc agatactgga aggattaaaa
     3361 taccttcacg acaatcagat agtccaccgg gatataaagg gtgacaatgt attgattaat
     3421 acctacagtg gtgtgctcaa gatctctgac ttcgggacat ccaagaggct tgctggcata
     3481 aacccatgta ccgaaacctt tacaggtacc cttcagtata tggcgccaga aataatcgat
     3541 aaaggaccac gaggctatgg aaaggccgcc gacatttggt cactgggctg cacaatcatc
     3601 gaaatggcta ctggaaaacc accattttat gaactaggag agccacaggc agccatgttc
     3661 aaggtgggga tgtttaaggt ccacccggag atcccagagt ccatgtcggc ggaggccaag
     3721 gcgttcatac tgaagtgttt tgaaccagac cctgacaaga gagcctgtgc taatgacttg
     3781 cttattgatg agttcttaaa agtgtccagc aaaaagaaaa agacacaacc caaactttca
     3841 gctctctcga ctggatcgaa tgagtatctt agaagcatct ccctgccggt ccctgtcctg
     3901 gtagaagaca ccagcagcag tagcgagtat ggctctgttt ctcctgatac agagctgaag
     3961 gcggacccct tctccttcaa agccagagct aagtcctgtg gagaaaagga cgggaaaggg
     4021 atacggacat tgttcctggg cattccagat gaaaattttg aagatcacag tgcgccccca
     4081 tctcctgaag agaaagactc gggctttttt atgctgagaa aggatagtga gaggcgagct
     4141 acccttcaca gaatcctgac cgaagaccag gacaaggttg taaggaacct aatggaatct
     4201 ctggcccagg gtgctgaaga gcctaaacta aagtgggaac acatcacaac cctcatctcg
     4261 agcctcagag agtttgtgag gtccactgac cgaaaaatca tagccactac actgtccaag
     4321 ctcaagctgg agctggactt cgacagccac ggcatcagcc aggtgcaggt ggtcctcttt
     4381 ggcttccagg atgcggtcaa taaagttctt cggaatcata acatcaagcc acactggatg
     4441 tttgccctgg acagcatcat ccgaaaggct gtgcagactg caattaccat tctggttcca
     4501 gagttgaggc cacattttag ccttgcatca gagagcgaca cagctgatcc ggaagacctg
     4561 gatgtcgaag atgaacacga ggaattgtct tcaaatcaga cagtccgacg gcctcaggcc
     4621 atcactgagg acgctgtggc tacctcgggg gtgagcacgc taagctccac tgtgtcccat
     4681 gactcccaga atgcacaccg gtcgctgaac gtgcagcttg gaaggatgaa gattgaaact
     4741 aataggttac ttgaagagct ggttcggaaa gagagagagt tacaggcact cctccatcaa
     4801 gccattgagg aaaaagacca agaaattagg cacctgaagc tcaagtccca gcccatagat
     4861 atccctgggt ttcctgtgtg ccacctgaac tctcctggca cgaccacgga agattctgaa
     4921 cttcctggct ggctgagaga aaatggagct gatgaagaca ctataagtcg gtttctggct
     4981 gaggattaca cgctagtgga tgttctttac tacgtcacac gtgatgatct gaagtgcctc
     5041 agactaaggg gcgggatgct gtgcacgctg tggaaggcca tcattgactt tcggaacaaa
     5101 tgcctcgagg actacaaaga ccatgacggt gattataaag atcatgacat cgactacaag
     5161 gatgacgatg acaagtagtg agggcccgtt taaacccgct gatcagcctc gactgtgcct
     5221 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     5281 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     5341 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     5401 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc
     5461 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     5521 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     5581 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     5641 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     5701 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     5761 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     5821 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     5881 gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt
     5941 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     6001 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     6061 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     6121 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     6181 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     6241 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcaagaga
     6301 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg
     6361 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg
     6421 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt
     6481 ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg gccacgacgg
     6541 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat
     6601 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat
     6661 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg
     6721 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg
     6781 atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc
     6841 tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc
     6901 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg
     6961 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg
     7021 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca
     7081 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac
     7141 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
     7201 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     7261 tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa
     7321 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     7381 tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta
     7441 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     7501 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     7561 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     7621 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     7681 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     7741 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     7801 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     7861 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     7921 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     7981 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     8041 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     8101 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     8161 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     8221 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     8281 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     8341 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggttt
     8401 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     8461 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     8521 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     8581 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     8641 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     8701 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     8761 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     8821 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     8881 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     8941 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     9001 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     9061 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     9121 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     9181 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     9241 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     9301 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     9361 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     9421 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     9481 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     9541 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     9601 gccacctgac gtc
//
