LOCUS       V041216                 8013 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V041216
VERSION     V041216
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..2693
                     /label="Hspa8(NM_031165)"
                     /note="Hspa8(NM_031165)"
                     /gene="Hspa8"
     CDS             2700..2765
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    2789..3377
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     polyA_signal    3443..3491
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal&nbsp;(Cole and Stacy, 1985)"
                     /label="HSV TK poly(A) signal"
     rep_origin      3693..4121
                     /direction=RIGHT
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     promoter        4135..4464
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             4531..5325
                     /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    5501..5622
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     primer_bind     complement(5671..5687)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    5695..5711
                     /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(5719..5749)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    5764..5785
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6073..6661)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(6832..7692)
                     /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(7693..7797)
                     /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 taagggacct gcagttggca
      781 ttgatctcgg caccacctac tcctgtgtgg gtgtcttcca gcatggaaag gtggaaatta
      841 ttgccaatga ccagggtaac cgcaccacgc caagctatgt tgctttcacg gacacagaga
      901 gattaattgg ggatgcggcc aagaatcagg ttgcaatgaa ccccaccaac acagtttttg
      961 atgccaaacg tctgatcggg cgtaggtttg atgatgctgt tgttcagtct gatatgaagc
     1021 actggccctt catggtggtg aatgatgcag gcaggcccaa ggtccaagtg gaatacaaag
     1081 gggagacaaa aagtttctac ccagaggaag tgtcctccat ggttctgaca aagatgaagg
     1141 aaattgcaga agcgtacctc ggaaagaccg ttaccaacgc tgtggtcaca gtgcccgctt
     1201 acttcaatga ctctcagcga caggcaacaa aagatgctgg aactattgct ggcctcaatg
     1261 tacttcgaat catcaatgaa ccaactgctg ctgctattgc ttacggctta gataagaagg
     1321 tcggagctga aaggaatgtg ctcatttttg acttgggagg tggcactttt gatgtgtcaa
     1381 tcctcactat tgaggatgga atttttgagg tcaaatcaac agctggagac acccacttag
     1441 gtggagaaga ttttgacaac cgaatggtca atcatttcat tgctgagttc aagcgaaagc
     1501 acaagaaaga catcagtgag aacaagagag ctgtccgccg tctccgcacg gcctgcgagc
     1561 gggccaagcg caccctctcc tccagcaccc aggccagtat tgagattgat tctctctatg
     1621 agggaattga cttctatacc tccattaccc gggctcgatt tgaggagttg aatgctgacc
     1681 tgttccgtgg cacactggac cctgtagaga aggcccttcg agatgccaag ctggacaagt
     1741 cacagatcca tgatattgtc ttggtgggtg gttctaccag aatccccaag attcagaaac
     1801 ttctgcaaga cttcttcaat ggaaaagagc tgaacaagag cattaacccc gatgaagctg
     1861 ttgcctatgg tgcagctgtc caggcagcca ttctatctgg agacaagtct gagaacgttc
     1921 aggatttgct gctcttggat gtcactcctc tttcccttgg tattgaaact gctggcggag
     1981 tcatgactgt cctcatcaag cgcaatacca ccatccccac caagcagaca cagactttca
     2041 ccacctactc tgacaaccag cctggtgtac tcattcaggt gtatgaaggt gaaagggcca
     2101 tgaccaagga caacaacctg cttggaaagt tcgagctcac aggcatccct ccagcacccc
     2161 gtggggtccc tcagattgag gttacttttg acatcgatgc caatggcatc ctcaatgttt
     2221 ctgctgtaga taagagcaca ggaaaggaga acaagatcac catcaccaat gacaagggcc
     2281 gcttgagtaa ggaagatatt gagcgcatgg tccaagaagc tgagaagtac aaggctgagg
     2341 atgagaagca gagagataag gtttcctcca agaactcact ggagtcctat gccttcaaca
     2401 tgaaagcaac tgtggaagat gagaaacttc aaggcaagat caatgatgag gacaaacaga
     2461 agattcttga caagtgcaat gaaatcatca gctggctgga taagaaccag actgcagaga
     2521 aggaagaatt tgagcatcag cagaaagaac tggagaaagt ctgcaaccct atcattacca
     2581 agctgtacca gagtgcaggt ggcatgcctg ggggaatgcc tggtggcttc ccaggtggag
     2641 gagctccccc atctggtggt gcttcttcag gccccaccat tgaagaggtg gatggatccg
     2701 actacaaaga ccatgacggt gattataaag atcatgacat cgactacaag gatgacgatg
     2761 acaagtagtg agggcccgat atctcgacaa tcaacctctg gattacaaaa tttgtgaaag
     2821 attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     2881 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     2941 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     3001 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     3061 ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     3121 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     3181 gaagctgacg tcctttccat ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac
     3241 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct
     3301 gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct
     3361 ttgggccgcc tccccgcctg gaaacggggg aggctaactg aaacacggaa ggagacaata
     3421 ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca cgggtgttgg
     3481 gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc
     3541 gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag
     3601 ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcaga
     3661 tctgcgcagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     3721 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     3781 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     3841 aaatcggggc atccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     3901 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     3961 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     4021 caaccctatc tcggtctatt cttttgattt ataagggatt ttggggattt cggcctattg
     4081 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt
     4141 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     4201 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     4261 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     4321 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     4381 tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt
     4441 tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct
     4501 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
     4561 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
     4621 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
     4681 accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg
     4741 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
     4801 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
     4861 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
     4921 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
     4981 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
     5041 ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
     5101 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
     5161 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
     5221 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
     5281 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
     5341 tcgcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc
     5401 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     5461 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     5521 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     5581 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     5641 gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     5701 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     5761 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     5821 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     5881 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     5941 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     6001 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     6061 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     6121 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     6181 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     6241 cccttcggga agcgtggcgc tttctcaatg ctcacgctgt aggtatctca gttcggtgta
     6301 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     6361 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     6421 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     6481 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     6541 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     6601 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     6661 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     6721 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     6781 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     6841 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     6901 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     6961 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     7021 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     7081 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     7141 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     7201 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     7261 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     7321 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     7381 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     7441 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     7501 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     7561 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     7621 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     7681 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     7741 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     7801 atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg
     7861 tcgactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct
     7921 tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc
     7981 ttgaccgaca attgcatgaa gaatctgctt agg
//
