LOCUS       V034554                 9851 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V034554
VERSION     V034554
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     polyA_signal    216..350
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     promoter        398..502
                     /gene="bla"
                     /label="AmpR promoter"
     CDS             503..1363
                     /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"
     rep_origin      1534..2119
                     /direction=RIGHT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     protein_bind    2407..2428
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     promoter        2443..2473
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    2481..2497
                     /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"
     primer_bind     2505..2521
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     LTR             2650..3283
                     /note="3' long terminal repeat (LTR) from HIV-1"
                     /label="3' LTR"
     misc_feature    3491..4079
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     CDS             complement(4093..4692)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /note="confers resistance to puromycin"
                     /product="puromycin N-acetyltransferase"
                     /transl_table=1
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA*"
                     /label="PuroR"
     promoter        complement(4701..5030)
                     /note="SV40 enhancer and early promoter"
                     /label="SV40 promoter"
     CDS             complement(5041..5787)
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to
                     promoters containing the tet operator in the presence of
                     doxycycline"
                     /transl_table=1
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG*"
                     /label="Tet-On® 3G"
     promoter        complement(5806..6316)
                     /note="human phosphoglycerate kinase 1 promoter"
                     /label="hPGK promoter"
     promoter        6333..6700
                     /note="3rd-generation Tet-responsive promoter that can be
                     activated by binding of Tet-On® 3G, modified to eliminate
                     binding sites for endogenous mammalian transcription
                     factors"
                     /label="TRE3GS promoter"
     CDS             6713..7276
                     /label="Fdx1(NM_007996)"
                     /note="Fdx1(NM_007996)"
                     /gene="Fdx1"
     CDS             7283..7348
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     polyA_signal    7531..7665
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     misc_feature    7709..7824
                     /note="central polypurine tract and central termination
                     sequence of HIV-1 (lacking the first T)"
                     /label="cPPT/CTS"
     CDS             complement(8087..8131)
                     /codon_start=1
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /product="antigenic peptide corresponding to amino acids
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik
                     et al., 2013)"
                     /transl_table=1
                     /translation="KNEQELLELDKWASL"
                     /label="gp41 peptide"
     misc_feature    8316..8549
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
                     /label="RRE"
     misc_feature    9046..9171
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
                     /label="HIV-1 Ψ"
     LTR             9218..9851
                     /note="3' long terminal repeat (LTR) from HIV-1"
                     /label="3' LTR"
ORIGIN
        1 actactaagt ttgtagtaca tatttaacaa atacaatttc tttaaaaact actaagtttg
       61 tagtacatat ttaacaaata caatttcttt aaaatgaaaa taattcagag gaatcacagg
      121 tttagagtaa atgaaaccac taggtaattg gcagtggtaa tagggtatgg ggtgggaagt
      181 ttgggatgat tttggttagc ttgagttatc cagttgatcc agacatgata agatacattg
      241 atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt
      301 gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt gatctcccga
      361 tccgtcgacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
      421 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
      481 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
      541 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
      601 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
      661 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
      721 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
      781 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
      841 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
      901 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
      961 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     1021 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     1081 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     1141 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     1201 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     1261 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     1321 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     1381 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     1441 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     1501 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     1561 ctgctgcttg caaacaaaaa aaccgctacc agcggtggtt tgtttgccgg atcaagagct
     1621 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     1681 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     1741 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     1801 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     1861 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     1921 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     1981 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     2041 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     2101 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     2161 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     2221 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     2281 agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc
     2341 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
     2401 cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
     2461 ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga
     2521 ccatgattac gccaagctct agctagaggt cgacggtaaa acgctagcaa tgtcaaggcc
     2581 tctcactctc tgatattcat ttctttgcaa gttataaata ctgaataata agatgacatg
     2641 aactactact gctagagatt ttccacactg actaaaaggg tctgagggat ctctagttac
     2701 cagagtcaca caacagacgg gcacacacta cttgaagcac tcaaggcaag ctttattgag
     2761 gcttaagcag tgggttccct agttagccag agagctccca ggctcagatc tggtctaacc
     2821 agagagaccc agtacaggca aaaagcagct gcttatatgc aggatctgag ggctcgccac
     2881 tccccagtcc cgcccaggcc acgcctccct ggaaagtccc cagcggaaag tcccttgtag
     2941 caagctcgat atcagcagtt cttgaagtac tccggatgca gctctcgggc cacgtgatga
     3001 aatgctaggc ggctgtcaaa cctccactct aacacttctc tctccgggtc atccatccca
     3061 tgcaggctca cagggtgtaa caagctggtg ttctctcctt tattggcctc ttctacctta
     3121 tctggctcaa ctggtactag cttgtagcac catccaaagg tcagtggata tctgacccct
     3181 ggccctggtg tgtagttctg ctaatcaggg aagtagcctt gtgtgtggta gatccacaga
     3241 tcaaggatat cttgtcttcg ttgggagtga attagccctt ccagtcccct cttttctttt
     3301 aaaaagtggc taagatctac agctgccttg taagtcattg gtcttaaagg tacctgaggt
     3361 gtgactggaa aaacccacct cctcctcctc ttgtgcttct agccaggcac aatcagcatt
     3421 ggtagctgct gtattgctac ttgtgattgc tccatgtttt tctaggtctc gactgcagaa
     3481 ttaattccag gcggggaggc ggcccaaagg gagatccgac tcgtctgagg gcgaaggcga
     3541 agacgcggaa gaggccgcag agccggcagc aggccgcggg aaggaaggtc cgctggattg
     3601 agggccgaag ggacgtagca gaaggacgtc ccgcgcagaa tccaggtggc aacacaggcg
     3661 agcagccatg gaaaggacgt cagcttcccc gacaacacca cggaattgtc agtgcccaac
     3721 agccgagccc ctgtccagca gcgggcaagg caggcggcga tgagttccgc cgtggcaata
     3781 gggaggggga aagcgaaagt cccggaaagg agctgacagg tggtggcaat gccccaacca
     3841 gtgggggttg cgtcagcaaa cacagtgcac accacgccac gttgcctgac aacgggccac
     3901 aactcctcat aaagagacag caaccaggat ttatacaagg aggagaaaat gaaagccata
     3961 cgggaagcaa tagcatgata caaaggcatt aaagcagcgt atccacatag cgtaaaagga
     4021 gcaacatagt taagaatacc agtcaatctt tcacaaattt tgtaatccag aggttgattg
     4081 ttccagacgc gttcaggcac cgggcttgcg ggtcatgcac caggtgcgcg gtccttcggg
     4141 cacctcgacg tcggcggtga cggtgaagcc gagccgctcg tagaagggga ggttgcgggg
     4201 cgcggaggtc tccaggaagg cgggcacccc ggcgcgctcg gccgcctcca ctccggggag
     4261 cacgacggcg ctgcccagac ccttgccctg gtggtcgggc gagacgccga cggtggccag
     4321 gaaccacgcg ggctccttgg gccggtgcgg cgccaggagg ccttccatct gttgctgcgc
     4381 ggccagccgg gaaccgctca actcggccat gcgcgggccg atctcggcga acaccgcccc
     4441 cgcttcgacg ctctccggcg tggtccagac cgccaccgcg gcgccgtcgt ccgcgaccca
     4501 caccttgccg atgtcgagcc cgacgcgcgt gaggaagagt tcttgcagct cggtgacccg
     4561 ctcgatgtgg cggtccgggt cgacggtgtg gcgcgtggcg gggtagtcgg cgaacgcggc
     4621 ggcgagggtg cgtacggccc gggggacgtc gtcgcgggtg gcgaggcgca ccgtgggctt
     4681 gtactcggtc atggtcgcgt tttgcaaaag cctaggcctc caaaaaagcc tcctcactac
     4741 ttctggaata gctcagaggc cgaggcggcc tcggcctctg cataaataaa aaaaattagt
     4801 cagccatggg gcggagaatg ggcggaactg ggcggagtta ggggcgggat gggcggagtt
     4861 aggggcggga ctatggttgc tgactaattg agatgcatgc tttgcatact tctgcctgct
     4921 ggggagcctg gggactttcc acacctggtt gctgactaat tgagatgcat gctttgcata
     4981 cttctgcctg ctggggagcc tggggacttt ccacacccta actgacacac attcgcgcgt
     5041 ttacccgggg agcatgtcaa ggtcaaaatc gtcaagagcg tcagcaggca gcatatcaag
     5101 gtcaaagtcg tcaagggcat cggctgggag catgtctaag tcaaaatcgt caagggcgtc
     5161 ggtcggcccg ccgctttcgc actttagctg tttctccagg ccacatatga ttagttccag
     5221 gccgaaaagg aaggcaggtt cggctccctg ccggtcgaac agctcaattg cttgtttcag
     5281 aagtgggggc atagaatcgg tggtaggtgt ctctctttcc tcttttgcta cttgatgctc
     5341 ctgttcctcc aatacgcagc ccagtgtaaa gtggcccacg gcggacagag cgtacagtgc
     5401 gttctccagg gagaagcctt gctgacacag gaacgcgagc tgattttcca gggtttcgta
     5461 ctgtttctct gttgggcggg tgccgagatg cactttagcc ccgtcgcgat gtgagaggag
     5521 agcacagcgg tatgacttgg cgttgttccg cagaaagtct tgccatgact cgccttccag
     5581 ggggcaggag tgggtatgat gcctgtccag catctcgatt ggcagggcat cgagcagggc
     5641 ccgcttgttc ttcacgtgcc agtacagggt aggctgctca actcccagct tttgagcgag
     5701 tttccttgtc gtcaggcctt cgataccgac tccattgagt aattccagag cagagtttat
     5761 gactttgctc ttgtccagtc tagacatggt aattcgatga tccccctggg gagagaggtc
     5821 ggtgattcgg tcaacgaggg agccgactgc cgacgtgcgc tccggaggct tgcagaatgc
     5881 ggaacaccgc gcgggcagga acagggccca cactaccgcc ccacaccccg cctcccgcac
     5941 cgccccttcc cggccgctgc tctcggcgcg ccctgctgag cagccgctat tggccacagc
     6001 ccatcgcggt cggcgcgctg ccattgctcc ctggcgctgt ccgtctgcga gggtactagt
     6061 gagacgtgcg gcttccgttt gtcacgtccg gcacgccgcg aaccgcaagg aaccttcccg
     6121 acttaggggc ggagcaggaa gcgtcgccgg ggggcccaca agggtagcgg cgaagatccg
     6181 ggtgacgctg cgaacggacg tgaagaatgt gcgagaccca gggtcggcgc cgctgcgttt
     6241 cccggaacca cgcccagagc agccgcgtcc ctgcgcaaac ccagggctgc cttggaaaag
     6301 gcgcaacccc aaccccgtgg aattatcacc tcgagtttac tccctatcag tgatagagaa
     6361 cgtatgaaga gtttactccc tatcagtgat agagaacgta tgcagacttt actccctatc
     6421 agtgatagag aacgtataag gagtttactc cctatcagtg atagagaacg tatgaccagt
     6481 ttactcccta tcagtgatag agaacgtatc tacagtttac tccctatcag tgatagagaa
     6541 cgtatatcca gtttactccc tatcagtgat agagaacgta taagctttgc ttatgtaaac
     6601 cagggcgcct ataaaagagt gctgattttt tgagtaaact tcaattccac aacacttttg
     6661 tcttatacca actttccgta ccacttccta ccctcgtaaa gaattcgcca ccatggcggc
     6721 cgctccgggc gcccgactcc tgcgcgcggc ctgcgcctcc gtccctttcc gcggccttga
     6781 ccgctgtcgg ctgctggtct gcgggaccgg agcgggaact gccatctctc cgtggacccc
     6841 gagtccccgc ctgcatgcag aggccgggcc cggccggccg ctgagcgtgt ctgcgcgcgc
     6901 gcggagcagc tcagaagata agataacagt ccacttcaag aaccgagatg gcgagacgct
     6961 aacgaccaag gggaaaattg gcgactctct gctagatgtt gtgattgaga acaacttaga
     7021 tatcgatggg tttggtgcgt gtgagggaac gttggcttgc tctacttgtc atcttatctt
     7081 tgaggatcac atctatgaga agttagatgc cattactgat gaagagaatg acatgcttga
     7141 cctggctttt ggactaacag acaggtcaag gttgggctgc caagtttgtc tgaccaaggc
     7201 tatggacaat atgactgtgc gtgtgcctga agcagtggcg gatgtccgac agtctgttga
     7261 catgagcaag aattccaccg gtgactacaa agaccatgac ggtgattata aagatcatga
     7321 catcgactac aaggatgacg atgacaagta ggatccagac cacctcccct gcgagctaag
     7381 ctggacagcc aatgacgggt aagagagtga catttttcac taacctaaga caggagggcc
     7441 gtcagagcta ctgcctaatc caaagacggg taaaagtgat aaaaatgtat cactccaacc
     7501 taagacaggc gcagcttccg agggatttga gatccagaca tgataagata cattgatgag
     7561 tttggacaaa ccaaaactag aatgcagtga aaaaaatgcc ttatttgtga aatttgtgat
     7621 gctattgcct tatttgtaac cattataagc tgcaataaac aagttaagtc gataaactgg
     7681 atctctgctg tccctgtaat aaacccgaaa attttgaatt tttgtaattt gtttttgtag
     7741 ttctttagtt tgtatgtctg ttgctattat gtctactatt ctttcccctg cactgtaccc
     7801 cccaatcccc ccttttcttt taaaggcgat accgtcgaga tccgttcact aatcgaatgg
     7861 atctgtctct gtctctctct ccaccttctt cttctattcc ttcgggcctg tcgggtcccc
     7921 tcggggttgg gaggtgggtc tgaaacgata atggtgaata tccctgccta actctattca
     7981 ctatagaaag tacagcaaaa actattctta aacctaccaa gcctcctact atcattatga
     8041 ataattttat ataccacagc caatttgtta tgttaaacca attccacaaa cttgcccatt
     8101 tatctaattc caataattct tgttcattct tttcttgctg gttttgcgat tcttcaatta
     8161 aggagtgtat taagcttgtg taattgttaa tttctctgtc ccactccatc caggtcgtgt
     8221 gattccaaat ctgttccaga gatttattac tccaactagc attccaaggc acagcagtgg
     8281 tgcaaatgag ttttccagag caaccccaaa tccccaggag ctgttgatcc tttaggtatc
     8341 tttccacagc caggattctt gcctggagct gcttgatgcc ccagactgtg agttgcaaca
     8401 gatgctgttg cgcctcaata gccctcagca aattgttctg ctgctgcact ataccagaca
     8461 ataattgtct ggcctgtacc gtcagcgtca ttgacgctgc gcccatagtg cttcctgctg
     8521 ctcccaagaa cccaaggaac aaagctccta ttcccactgc tcttttttct ctctgcacca
     8581 ctcttctctt tgccttggtg ggtgctactc ctaatggttc aatttttact actttatatt
     8641 tatataattc acttctccaa ttgtccctca tatctcctcc tccaggtctg aagatcagcg
     8701 gccggccgct tgctgtgcgg tggtcttact tttgttttgc tcttcctcta tcttgtctaa
     8761 agcttccttg gtgtctttta tctctatcct ttgatgcaca caatagaggg ttgctactgt
     8821 attatataat gatctaagtt cttctgatcc tgtctgaagg gatggttgta gctgtcccag
     8881 tatttgtcta cagccttctg atgtttctaa caggccagga ttaactgcga atcgttctag
     8941 ctccctgctt gcccatacta tatgttttaa tttatatttt ttctttcccc ctggccttaa
     9001 ccgaattttt tcccatcgcg atctaattct cccccgctta atactgacgc tctcgcaccc
     9061 atctctctcc ttctagcctc cgctagtcaa aatttttggc gtactcacca gtcgccgccc
     9121 ctcgcctctt gccgtgcgcg cttcagcaag ccgagtcctg cgtcgagaga gctcctctgg
     9181 tttccctttc gctttcaagt ccctgttcgg gcgccactgc tagagatttt ccacactgac
     9241 taaaagggtc tgagggatct ctagttacca gagtcacaca acagacgggc acacactact
     9301 tgaagcactc aaggcaagct ttattgaggc ttaagcagtg ggttccctag ttagccagag
     9361 agctcccagg ctcagatctg gtctaaccag agagacccag tacaggcaaa aagcagctgc
     9421 ttatatgcag gatctgaggg ctcgccactc cccagtcccg cccaggccac gcctccctgg
     9481 aaagtcccca gcggaaagtc ccttgtagca agctcgatat cagcagttct tgaagtactc
     9541 cggatgcagc tctcgggcca cgtgatgaaa tgctaggcgg ctgtcaaacc tccactctaa
     9601 cacttctctc tccgggtcat ccatcccatg caggctcaca gggtgtaaca agctggtgtt
     9661 ctctccttta ttggcctctt ctaccttatc tggctcaact ggtactagct tgtagcacca
     9721 tccaaaggtc agtggatatc tgacccctgg ccctggtgtg tagttctgct aatcagggaa
     9781 gtagccttgt gtgtggtaga tccacagatc aaggatatct tgtcttcttt gggagtgaat
     9841 tagcccttcc a
//
