LOCUS       V044322                 6996 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V044322
VERSION     V044322
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     enhancer        50..429
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        430..633
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     promoter        678..696
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /label="T7 promoter"
     CDS             756..1676
                     /label="Rrad(NM_019662)"
                     /note="Rrad(NM_019662)"
                     /gene="Rrad"
     CDS             1683..1748
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    1772..2360
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    2426..2474
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      2676..3104
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        3118..3447
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             3514..4308
                     /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    4484..4605
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(4654..4670)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    4678..4694
                     /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(4702..4732)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    4747..4768
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(5056..5644)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(5815..6675)
                     /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(6676..6780)
                     /gene="bla"
                     /label="AmpR promoter"
ORIGIN
        1 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
       61 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      121 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      181 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      241 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      301 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      361 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      421 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      481 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      541 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      601 cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga acccactgct
      661 tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg ctagcgttta
      721 aacttaagct tggtaccgag ctcgaattcg ccaccatgac cctgaatggt ggcagcggag
      781 caagcgggag ccgtggtgcg ggccgggagc gcgaccgccg tcgaggcagc accccatggg
      841 gcccggcacc cccactgcac cgcaggagca tgccggtgga cgagcgagac cttcaggcgg
      901 cgctagcccc gggctccctg gcaacgactg ccgctggaac tcggacacag ggccagaggc
      961 tggactggcc cgaaggctcc tccgactcgc tcagttctgg aggcagtggg tcagaggagg
     1021 gcgtttacaa ggtgctgctg ctcggggcgc ctggtgtggg caaaagcgct ctggcgcgca
     1081 tctttggtgg tatagaagac ggacctgaag cagaagcagc agggcacaca tatgaccgtt
     1141 ctatcactgt ggatggagaa gaagcgtcac tattggtcta tgacatttgg gaacaggatg
     1201 ggggctgctg gctgcctggc cactgcatgg ccatggggga tgcatacgtc atcgtgtact
     1261 ccatcacgga caagggcagc tttgagaaag cctcagaact ccgggtccag ctgcggcggg
     1321 cccggcagac agacgacgtg cccatcattc tcgtgggcaa caagagtgac ctggtgcgct
     1381 ctcgtgaggt ctctgtggat gagggccggg cctgcgcagt ggtcttcgac tgcaagttca
     1441 tcgagacatc agcagcactg catcacaatg tccaggcact attcgagggg gttgtgcgcc
     1501 agatacgcct gcgcagggac agcaaagaag ataacgcccg gcggcaagct ggcactcggc
     1561 gacgggagag ccttggcaag aaggccaaac gctttctggg ccgcatcgtg gctcgcaaca
     1621 gccgcaagat ggccttccgc gccaagtcca agtcctgcca tgacctctct gttctcggat
     1681 ccgactacaa agaccatgac ggtgattata aagatcatga catcgactac aaggatgacg
     1741 atgacaagta gtgagggccc gatatctcga caatcaacct ctggattaca aaatttgtga
     1801 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     1861 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     1921 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     1981 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     2041 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     2101 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     2161 ggggaagctg acgtcctttc catggctgct cgcctgtgtt gccacctgga ttctgcgcgg
     2221 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     2281 gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc
     2341 cctttgggcc gcctccccgc ctggaaacgg gggaggctaa ctgaaacacg gaaggagaca
     2401 ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacgggtgt
     2461 tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc
     2521 accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc
     2581 aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc
     2641 agatctgcgc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     2701 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     2761 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     2821 tctaaatcgg ggcatccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     2881 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     2941 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     3001 actcaaccct atctcggtct attcttttga tttataaggg attttgggga tttcggccta
     3061 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
     3121 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     3181 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     3241 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     3301 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     3361 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
     3421 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
     3481 tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     3541 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     3601 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     3661 aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg
     3721 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     3781 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     3841 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     3901 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     3961 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     4021 ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc
     4081 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     4141 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     4201 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     4261 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     4321 ggttcgcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     4381 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     4441 cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc
     4501 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     4561 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc
     4621 gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     4681 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     4741 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     4801 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4861 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     4921 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     4981 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     5041 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     5101 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     5161 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     5221 tctcccttcg ggaagcgtgg cgctttctca atgctcacgc tgtaggtatc tcagttcggt
     5281 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     5341 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     5401 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     5461 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     5521 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     5581 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     5641 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5701 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     5761 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     5821 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     5881 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     5941 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     6001 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     6061 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     6121 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     6181 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     6241 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     6301 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     6361 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     6421 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     6481 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     6541 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     6601 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     6661 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6721 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     6781 cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta
     6841 tggtcgactc tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct
     6901 gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa
     6961 ggcttgaccg acaattgcat gaagaatctg cttagg
//
