LOCUS       V025333                 9326 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V025333
VERSION     V025333
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     LTR             234..414
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
                     /label="5' LTR (truncated)"
     misc_feature    458..583
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
                     /label="HIV-1 Ψ"
     misc_feature    1076..1309
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
                     /label="RRE"
     CDS             1493..1537
                     /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    1804..1921
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
                     /label="cPPT/CTS"
     promoter        2011..2214
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     CDS             2317..3369
                     /label="WNT4(NM_030761)"
                     /note="WNT4(NM_030761)"
                     /gene="WNT4"
     CDS             3376..3441
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3485..3696
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3709..3977
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      4006..4015
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             4036..4698
                     /codon_start=1
                     /product="green fluorescent protein 2 from Pontellina
                     plumata, also known as ppluGFP2 (Shagin et al., 2004)"
                     /transl_table=1
                     /translation="PAMEIECRITGTLNGVEFELVGGGEGTPKQGRMTNKMKSTKGALT
                     FSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYRY
                     EAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDGG
                     YYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA"
                     /label="CopGFP"
     CDS             4768..4821
                     /codon_start=1
                     /note="Eukaryotic ribosomes fail to insert a peptide bond
                     between the Gly and Pro residues, yielding separate
                     polypeptides."
                     /product="2A peptide from&nbsp;Thosea asigna virus capsid
                     protein"
                     /protein_id=""
                     /transl_table=1
                     /translation="EGRGSLLTCGDVEENPGP"
                     /label="T2A"
     CDS             4822..5421
                     /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"
     misc_feature    5422..6009
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6083..6316
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6388..6509
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6528..6663
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6701..6717)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6725..6741
                     /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(6749..6779)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6794..6815
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7103..7691)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7862..8722)
                     /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(8723..8827)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9301..9317
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 fwd"
ORIGIN
        1 acgcgtgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga tgagttagca
       61 acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag taaggtggta
      121 cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac gaaccactga
      181 attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacaata aacgggtctc
      241 tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta
      301 agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact
      361 ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtggcg
      421 cccgaacagg gacctgaaag cgaaagggaa accagagctc tctcgacgca ggactcggct
      481 tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt
      541 gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag
      601 aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt
      661 aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt
      721 agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg
      781 atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag
      841 gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag
      901 taagaccacc gcacagcaag cggccactga tcttcagacc tggaggagga gatatgaggg
      961 acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag
     1021 cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag
     1081 ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcc tcaatgacgc
     1141 tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga
     1201 gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc
     1261 aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg
     1321 gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata
     1381 aatctctgga acagattgga atcacacgac ctggatggag tgggacagag aaattaacaa
     1441 ttacacaagc ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga
     1501 acaagaatta ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa
     1561 ttggctgtgg tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat
     1621 agtttttgct gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt
     1681 tcagacccac ctcccaaccc cgaggggacc cgacaggccc gaaggaatag aagaagaagg
     1741 tggagagaga gacagagaca gatccattcg attagtgaac ggatctcgac ggtatcggtt
     1801 aacttttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
     1861 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
     1921 tatcgatact agtattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     1981 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     2041 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     2101 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     2161 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggttt atataagcag agctcgttta
     2221 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagattc
     2281 tagagctagc ctgatacgaa ctcggaattc gccaccatga gtccccgctc gtgcctgcgt
     2341 tcgctgcgcc tcctcgtctt cgccgtcttc tcagccgccg cgagcaactg gctgtacctg
     2401 gccaagctgt cgtcggtggg gagcatctca gaggaggaga cgtgcgagaa actcaagggc
     2461 ctgatccaga ggcaggtgca gatgtgcaag cggaacctgg aagtcatgga ctcggtgcgc
     2521 cgcggtgccc agctggccat tgaggagtgc cagtaccagt tccggaaccg gcgctggaac
     2581 tgctccacac tcgactcctt gcccgtcttc ggcaaggtgg tgacgcaagg gactcgggag
     2641 gcggccttcg tgtacgccat ctcttcggca ggtgtggcct ttgcagtgac gcgggcgtgc
     2701 agcagtgggg agctggagaa gtgcggctgt gacaggacag tgcatggggt cagcccacag
     2761 ggcttccagt ggtcaggatg ctctgacaac atcgcctacg gtgtggcctt ctcacagtcg
     2821 tttgtggatg tgcgggagag aagcaagggg gcctcgtcca gcagagccct catgaacctc
     2881 cacaacaatg aggccggcag gaaggccatc ctgacacaca tgcgggtgga atgcaagtgc
     2941 cacggggtgt caggctcctg tgaggtaaag acgtgctggc gagccgtgcc gcccttccgc
     3001 caggtgggtc acgcactgaa ggagaagttt gatggtgcca ctgaggtgga gccacgccgc
     3061 gtgggctcct ccagggcact ggtgccacgc aacgcacagt tcaagccgca cacagatgag
     3121 gacctggtgt acttggagcc tagccccgac ttctgtgagc aggacatgcg cagcggcgtg
     3181 ctgggcacga ggggccgcac atgcaacaag acgtccaagg ccatcgacgg ctgtgagctg
     3241 ctgtgctgtg gccgcggctt ccacacggcg caggtggagc tggctgaacg ctgcagctgc
     3301 aaattccact ggtgctgctt cgtcaagtgc cggcagtgcc agcggctcgt ggagttgcac
     3361 acgtgccgag gatccgacta caaagaccat gacggtgatt ataaagatca tgacatcgac
     3421 tacaaggatg acgatgacaa gtagcggccg cgaaggatct gcgatcgctc cggtgcccgt
     3481 cagtgggcag agcgcacatc gcccacagtc cccgagaagt tggggggagg ggtcggcaat
     3541 tgaacgggtg cctagagaag gtggcgcggg gtaaactggg aaagtgatgt cgtgtactgg
     3601 ctccgccttt ttcccgaggg tgggggagaa ccgtatataa gtgcagtagt cgccgtgaac
     3661 gttctttttc gcaacgggtt tgccgccaga acacagctga agcttcgagg ggctcgcatc
     3721 tctccttcac gcgcccgccg ccctacctga ggccgccatc cacgccggtt gagtcgcgtt
     3781 ctgccgcctc ccgcctgtgg tgcctcctga actgcgtccg ccgtctaggt aagtttaaag
     3841 ctcaggtcga gaccgggcct ttgtccggcg ctcccttgga gcctacctag actcagccgg
     3901 ctctccacgc tttgcctgac cctgcttgct caactctacg tctttgtttc gttttctgtt
     3961 ctgcgccgtt acagatccaa gctgtgaccg gcgcctacgc tagacgccac catggagagc
     4021 gacgagagcg gcctgcccgc catggagatc gagtgccgca tcaccggcac cctgaacggc
     4081 gtggagttcg agctggtggg cggcggagag ggcaccccca agcagggccg catgaccaac
     4141 aagatgaaga gcaccaaagg cgccctgacc ttcagcccct acctgctgag ccacgtgatg
     4201 ggctacggct tctaccactt cggcacctac cccagcggct acgagaaccc cttcctgcac
     4261 gccatcaaca acggcggcta caccaacacc cgcatcgaga agtacgagga cggcggcgtg
     4321 ctgcacgtga gcttcagcta ccgctacgag gccggccgcg tgatcggcga cttcaaggtg
     4381 gtgggcaccg gcttccccga ggacagcgtg atcttcaccg acaagatcat ccgcagcaac
     4441 gccaccgtgg agcacctgca ccccatgggc gataacgtgc tggtgggcag cttcgcccgc
     4501 accttcagcc tgcgcgacgg cggctactac agcttcgtgg tggacagcca catgcacttc
     4561 aagagcgcca tccaccccag catcctgcag aacgggggcc ccatgttcgc cttccgccgc
     4621 gtggaggagc tgcacagcaa caccgagctg ggcatcgtgg agtaccagca cgccttcaag
     4681 acccccatcg ccttcgccag atcccgcgct cagtcgtcca attctgccgt ggacggcacc
     4741 gccggacccg gctccaccgg atctcgcgag ggcagaggaa gtcttctaac atgcggtgac
     4801 gtggaggaga atcccggccc tatgaccgag tacaagccca cggtgcgcct cgccacccgc
     4861 gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta ccccgccacg
     4921 cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca agaactcttc
     4981 ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg
     5041 gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg
     5101 cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga aggcctcctg
     5161 gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac
     5221 caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc
     5281 gccggggtgc ccgccttcct ggagacctcc gcgccccgca acctcccctt ctacgagcgg
     5341 ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg
     5401 acccgcaagc ccggtgcctg aaatcaacct ctggattaca aaatttgtga aagattgact
     5461 ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg
     5521 tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg
     5581 ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg
     5641 tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct cctttccggg
     5701 actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc
     5761 tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaagctg
     5821 acgtcctttc catggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc
     5881 tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct
     5941 ctgcggcctc ttccgcgtct ccgccttcgc cctcagacga gtcggatctc cctttggccg
     6001 cctccccgcc tggtaccttt aagaccaatg acttacaagg cagctgtaga tcttagccac
     6061 tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaaa ataagatctg
     6121 ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     6181 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     6241 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     6301 tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt ataacttgca
     6361 aagaaatgaa tatcagagag tgagaggaac ttgtttattg cagcttataa tggttacaaa
     6421 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     6481 ggtttgtcca aactcatcaa tgtatcttat catgtctggc tctagctatc ccgcccctaa
     6541 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     6601 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     6661 gcctagactt ttgcagagac ggcccaaatt cgtaatcatg gtcatagctg tttcctgtgt
     6721 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     6781 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     6841 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     6901 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     6961 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     7021 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     7081 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     7141 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     7201 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     7261 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     7321 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     7381 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     7441 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     7501 cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct
     7561 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     7621 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     7681 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     7741 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     7801 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     7861 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     7921 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     7981 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     8041 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     8101 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     8161 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     8221 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     8281 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     8341 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     8401 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     8461 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     8521 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     8581 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     8641 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     8701 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     8761 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     8821 ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga
     8881 cattaaccta taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg
     8941 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
     9001 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct
     9061 ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa
     9121 taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc
     9181 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
     9241 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
     9301 gtaaaacgac ggccagtgcc aagctg
//
