LOCUS       V035087                 8202 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V035087
VERSION     V035087
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..2882
                     /label="KLHL5(NM_015990)"
                     /note="KLHL5(NM_015990)"
                     /gene="KLHL5"
     CDS             2889..2954
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    2978..3566
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    3632..3680
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      3882..4310
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        4324..4653
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             4720..5514
                     /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    5690..5811
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(5860..5876)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5884..5900
                     /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(5908..5938)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    5953..5974
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6262..6850)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7021..7881)
                     /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(7882..7986)
                     /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 ccaccatgtc tggttctcgt aaagagtttg
      781 atgtgaaaca gattttgaaa atcagatgga ggtggtttgg tcatcaagca tcatctccta
      841 attctacagt tgacagccag cagggagaat tttggaaccg aggacagact ggagcaaacg
      901 gtgggagaaa gtttttagat ccatgtagcc tacaattgcc tttggcttca attggttacc
      961 gaaggtccag ccaactggat tttcagaatt caccttcttg gccaatggca tccacctctg
     1021 aagtccctgc atttgagttt acagcagaag attgtggcgg tgcacattgg ctggatagac
     1081 cagaagtgga tgatggcact agtgaagaag aaaatgaatc tgattccagt tcatgcagga
     1141 cttccaatag tagtcagaca ttatcatcct gtcatactat ggagccatgt acatcagatg
     1201 aatttttcca agcccttaat catgccgagc aaacatttaa aaaaatggaa aactatttga
     1261 gacataaaca gttgtgtgat gtaattttag tcgctggtga tcgcagaatt ccagctcaca
     1321 gattggtgct ctcctctgtc tcagactatt ttgctgccat gtttactaat gatgtcagag
     1381 aggcaagaca agaagaaata aaaatggaag gtgtagaacc aaattcgttg tggtccttga
     1441 tccagtatgc ttatacaggc cgccttgaat taaaagaaga taatattgag tgcctgttat
     1501 ctacagcttg ccttcttcag ctttcacagg ttgtagaagc atgctgtaag tttttaatga
     1561 aacagcttca tccatccaac tgtcttggaa ttcgttcttt tgctgatgcc caaggttgta
     1621 cagatttgca taaagtggct cacaattata ctatggagca tttcatggaa gtaatcagaa
     1681 accaggaatt tgtattatta ccagccagcg aaattgcaaa gctcttggct agtgatgaca
     1741 tgaacattcc taatgaggag acaatattga atgcacttct tacttgggtc cgtcatgatt
     1801 tggaacagag acggaaagat ctaagtaaac ttttggctta tattaggcta cctcttcttg
     1861 caccacagtt cctggcagac atggaaaata atgtactttt tcgggatgat atagaatgtc
     1921 agaaactcat tatggaagca atgaagtacc atttattacc agagagacga cccatgttac
     1981 aaagtcctcg gacaaaacct aggaagtcaa ctgttggtac attatttgca gttgggggaa
     2041 tggattcaac aaaaggagca acaagcattg aaaagtatga tctccgtaca aatatgtgga
     2101 ctccagtagc aaatatgaat gggaggaggc tacagttcgg tgttgcagtg ctagatgaca
     2161 aactgtatgt ggttggagga agagatggac tgaagacttt gaatactgta gagtgctaca
     2221 accccaaaac aaaaacttgg agtgtgatgc cacctatgtc cacacataga catggccttg
     2281 gtgtggctgt actggaaggt cccatgtatg ccgtaggagg acatgatggc tggagctatc
     2341 tgaacacagt ggaaagatgg gaccctcagg ctcgccagtg gaattttgtt gccactatgt
     2401 ctacccctag gagtacagta ggtgtggcag tactaagtgg aaaactttat gcagttggtg
     2461 gtcgtgatgg aagttcttgt ctcaaatcag tagaatgttt tgatcctcat actaataagt
     2521 ggacactgtg tgcacagatg tcaaaaagga gaggtggcgt aggagtgacg acctggaatg
     2581 gactgctgta tgctataggg gggcacgatg ctcccgcatc caacttgact tccagactct
     2641 cagactgtgt ggaaagatat gatcccaaaa cagacatgtg gactgcagta gcatccatga
     2701 gcatcagcag agatgcagtg ggggtctgtt tacttggtga taagttatat gctgttgggg
     2761 ggtatgatgg acaggcatac cttaatactg tggaggctta tgatccccag acaaatgagt
     2821 ggacccaggt tgctccactg tgcctaggaa gagctggagc ttgtgttgtg actgtaaaat
     2881 taggatccga ctacaaagac catgacggtg attataaaga tcatgacatc gactacaagg
     2941 atgacgatga caagtagtga gggcccgata tctcgacaat caacctctgg attacaaaat
     3001 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     3061 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     3121 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     3181 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     3241 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     3301 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     3361 gttgtcgggg aagctgacgt cctttccatg gctgctcgcc tgtgttgcca cctggattct
     3421 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     3481 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     3541 gatctccctt tgggccgcct ccccgcctgg aaacggggga ggctaactga aacacggaag
     3601 gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa taaaacgcac
     3661 gggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga
     3721 taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca
     3781 ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc
     3841 catagcagat ctgcgcagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     3901 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     3961 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     4021 tcaagctcta aatcggggca tccctttagg gttccgattt agtgctttac ggcacctcga
     4081 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     4141 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     4201 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tggggatttc
     4261 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     4321 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     4381 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     4441 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     4501 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     4561 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     4621 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     4681 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     4741 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     4801 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     4861 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     4921 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     4981 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     5041 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     5101 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     5161 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     5221 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     5281 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     5341 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     5401 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     5461 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     5521 ctctggggtt cgcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga
     5581 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     5641 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     5701 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     5761 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     5821 tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     5881 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     5941 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     6001 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     6061 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     6121 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     6181 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     6241 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     6301 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     6361 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     6421 cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc tcacgctgta ggtatctcag
     6481 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     6541 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     6601 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     6661 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     6721 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     6781 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     6841 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     6901 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     6961 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     7021 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     7081 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     7141 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     7201 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     7261 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     7321 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     7381 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     7441 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     7501 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     7561 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     7621 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     7681 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     7741 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     7801 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     7861 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     7921 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     7981 tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat
     8041 cccctatggt cgactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg
     8101 ctccctgctt gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca
     8161 aggcaaggct tgaccgacaa ttgcatgaag aatctgctta gg
//
