LOCUS       V045319                 7062 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V045319
VERSION     V045319
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..1742
                     /label="Ctsk(NM_007802)"
                     /note="Ctsk(NM_007802)"
                     /gene="Ctsk"
     CDS             1749..1814
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    1838..2426
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    2492..2540
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      2742..3170
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        3184..3513
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             3580..4374
                     /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    4550..4671
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(4720..4736)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    4744..4760
                     /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(4768..4798)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    4813..4834
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(5122..5710)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(5881..6741)
                     /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(6742..6846)
                     /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 ccaccatgtg ggtgttcaag tttctgctgc
      781 tacccatggt gagctttgct ctgtctccgg aggaaatgct ggacacccag tgggagctat
      841 ggaagaagac tcaccagaag cagtataaca gcaaggtgga tgaaatctct cggcgtttaa
      901 tttgggagaa aaacctgaag caaatctctg cccataacct ggaggcctct cttggtgtcc
      961 atacatatga actggccatg aaccacttgg gagacatgac cagtgaagaa gtggttcaga
     1021 agatgacggg actcagaata cctccctctc gatcctacag taatgacact ctctataccc
     1081 cagagtggga aggcagggtc ccagactcca tcgactatcg aaagaaagga tacgttactc
     1141 cagtcaagaa ccagggccag tgtggttcct gttgggcttt cagctctgcc ggggccctgg
     1201 agggccaact caagaagaaa actggtaaac tcttagctct gagtccccag aatcttgtgg
     1261 actgtgtgac tgagaattat ggctgtggag gcggctatat gaccactgcc ttccaatacg
     1321 tgcagcagaa cggaggcatt gactctgaag atgcttaccc atatgtgggc caggatgaaa
     1381 gttgtatgta taacgccacg gcaaaggcag ctaaatgcag agggtacaga gagattcctg
     1441 tggggaacga gaaagccctg aagagagcag tggcgcgggt aggacccatc tctgtgtcca
     1501 tcgatgcaag cttggcatct ttccagtttt acagcagagg tgtgtactat gatgaaaatt
     1561 gtgaccgtga taatgtgaac catgcagtgt tggtggtggg ctatggcacc cagaagggaa
     1621 gcaagcactg gataattaaa aacagctggg gagagagctg gggaaacaaa ggatatgctc
     1681 tcttggctcg gaataagaac aacgcctgcg gcattaccaa catggccagc ttccccaaga
     1741 tgggatccga ctacaaagac catgacggtg attataaaga tcatgacatc gactacaagg
     1801 atgacgatga caagtagtga gggcccgata tctcgacaat caacctctgg attacaaaat
     1861 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     1921 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     1981 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     2041 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     2101 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     2161 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     2221 gttgtcgggg aagctgacgt cctttccatg gctgctcgcc tgtgttgcca cctggattct
     2281 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     2341 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     2401 gatctccctt tgggccgcct ccccgcctgg aaacggggga ggctaactga aacacggaag
     2461 gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa taaaacgcac
     2521 gggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga
     2581 taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca
     2641 ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc
     2701 catagcagat ctgcgcagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     2761 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     2821 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     2881 tcaagctcta aatcggggca tccctttagg gttccgattt agtgctttac ggcacctcga
     2941 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     3001 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     3061 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tggggatttc
     3121 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     3181 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     3241 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     3301 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     3361 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3421 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     3481 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     3541 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     3601 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     3661 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     3721 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     3781 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     3841 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     3901 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     3961 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     4021 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     4081 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     4141 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     4201 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     4261 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     4321 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     4381 ctctggggtt cgcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga
     4441 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     4501 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     4561 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     4621 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     4681 tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     4741 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     4801 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     4861 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     4921 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     4981 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5041 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     5101 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     5161 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     5221 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     5281 cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc tcacgctgta ggtatctcag
     5341 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     5401 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     5461 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     5521 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     5581 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     5641 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     5701 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     5761 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     5821 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     5881 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     5941 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6001 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     6061 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     6121 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     6181 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     6241 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     6301 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     6361 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     6421 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     6481 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     6541 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     6601 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     6661 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     6721 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     6781 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     6841 tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat
     6901 cccctatggt cgactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg
     6961 ctccctgctt gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca
     7021 aggcaaggct tgaccgacaa ttgcatgaag aatctgctta gg
//
