LOCUS       V035476                 9080 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V035476
VERSION     V035476
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             2303..2917
                     /label="PMF1(NM_007221)"
                     /note="PMF1(NM_007221)"
                     /gene="PMF1"
     CDS             2924..2989
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3033..3244
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3257..3525
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      3554..3563
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             3584..4246
                     /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             4316..4369
                     /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             4376..5170
                     /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="MAEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*"
                     /label="NeoR/KanR"
     misc_feature    5177..5765
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5837..6070
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6142..6263
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6282..6417
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6455..6471)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6479..6495
                     /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(6503..6533)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6548..6569
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6857..7445)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7616..8476)
                     /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(8477..8581)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9055..9071
                     /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 gaattcgcca ccatggccga agcaagtagc gccaatctag gcagcggctg
     2341 tgaggaaaaa aggcatgagg ggtcgtcttc ggaatctgtg ccacccggca ctaccatttc
     2401 gagggtgaag ctcctcgaca ccatggtgga cacttttctt cagaagctgg tcgccgccgg
     2461 cagctaccag agattcactg actgctataa gtgcttctac cagttgcagc ctgcgatgac
     2521 acagcaaatc tatgacaagt ttatagctca gttgcagaca tctatccggg aggaaatctc
     2581 tgacatcaaa gaggagggga acctagaagc tgtcttgaat gccttggata aaattgtgga
     2641 agaaggcaaa gtccgcaaag agccagcctg gcgccccagc gggatcccag agaaggatct
     2701 gcacagtgtt atggcaccct acttcctgca gcaacgggac accctgcggc gccatgtgca
     2761 gaaacaggag gccgagaacc agcagctggc agatgccgtc ctggcagggc ggaggcaggt
     2821 ggaggagctg cagctacagg tccaggccca gcagcaggcc tggcaggctc tacacagaga
     2881 acagagggag ctggttgctg tgctgaggga gcctgaggga tccgactaca aagaccatga
     2941 cggtgattat aaagatcatg acatcgacta caaggatgac gatgacaagt agcggccgcg
     3001 aaggatctgc gatcgctccg gtgcccgtca gtgggcagag cgcacatcgc ccacagtccc
     3061 cgagaagttg gggggagggg tcggcaattg aacgggtgcc tagagaaggt ggcgcggggt
     3121 aaactgggaa agtgatgtcg tgtactggct ccgccttttt cccgagggtg ggggagaacc
     3181 gtatataagt gcagtagtcg ccgtgaacgt tctttttcgc aacgggtttg ccgccagaac
     3241 acagctgaag cttcgagggg ctcgcatctc tccttcacgc gcccgccgcc ctacctgagg
     3301 ccgccatcca cgccggttga gtcgcgttct gccgcctccc gcctgtggtg cctcctgaac
     3361 tgcgtccgcc gtctaggtaa gtttaaagct caggtcgaga ccgggccttt gtccggcgct
     3421 cccttggagc ctacctagac tcagccggct ctccacgctt tgcctgaccc tgcttgctca
     3481 actctacgtc tttgtttcgt tttctgttct gcgccgttac agatccaagc tgtgaccggc
     3541 gcctacgcta gacgccacca tggagagcga cgagagcggc ctgcccgcca tggagatcga
     3601 gtgccgcatc accggcaccc tgaacggcgt ggagttcgag ctggtgggcg gcggagaggg
     3661 cacccccaag cagggccgca tgaccaacaa gatgaagagc accaaaggcg ccctgacctt
     3721 cagcccctac ctgctgagcc acgtgatggg ctacggcttc taccacttcg gcacctaccc
     3781 cagcggctac gagaacccct tcctgcacgc catcaacaac ggcggctaca ccaacacccg
     3841 catcgagaag tacgaggacg gcggcgtgct gcacgtgagc ttcagctacc gctacgaggc
     3901 cggccgcgtg atcggcgact tcaaggtggt gggcaccggc ttccccgagg acagcgtgat
     3961 cttcaccgac aagatcatcc gcagcaacgc caccgtggag cacctgcacc ccatgggcga
     4021 taacgtgctg gtgggcagct tcgcccgcac cttcagcctg cgcgacggcg gctactacag
     4081 cttcgtggtg gacagccaca tgcacttcaa gagcgccatc caccccagca tcctgcagaa
     4141 cgggggcccc atgttcgcct tccgccgcgt ggaggagctg cacagcaaca ccgagctggg
     4201 catcgtggag taccagcacg ccttcaagac ccccatcgcc ttcgccagat cccgcgctca
     4261 gtcgtccaat tctgccgtgg acggcaccgc cggacccggc tccaccggat ctcgcgaggg
     4321 cagaggaagt cttctaacat gcggtgacgt ggaggagaat cccggccctg ccaccatggc
     4381 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     4441 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     4501 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     4561 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     4621 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     4681 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     4741 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     4801 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     4861 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     4921 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     4981 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     5041 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     5101 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     5161 gttcttctga gtcgacaatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat
     5221 tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca
     5281 tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc
     5341 tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc
     5401 tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt
     5461 cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg
     5521 gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc
     5581 ctttccttgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta
     5641 cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg
     5701 gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc
     5761 cccgcggtac ctttaagacc aatgacttac aaggcagctg tagatcttag ccacttttta
     5821 aaagaaaagg ggggactgga agggctaatt cactcccaac gaagataaga tctgcttttt
     5881 gcttgtactg ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta
     5941 gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc
     6001 cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa
     6061 atctctagca gtagtagttc atgtcatctt attattcagt atttataact tgcaaagaaa
     6121 tgaatatcag agagtgagag gaacttgttt attgcagctt ataatggtta caaataaagc
     6181 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     6241 tccaaactca tcaatgtatc ttatcatgtc tggctctagc tatcccgccc ctaactccgc
     6301 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     6361 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     6421 acttttgcag agacggccca aattcgtaat catggtcata gctgtttcct gtgtgaaatt
     6481 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     6541 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     6601 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     6661 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     6721 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     6781 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     6841 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     6901 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     6961 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     7021 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     7081 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     7141 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     7201 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     7261 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     7321 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     7381 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     7441 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     7501 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     7561 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     7621 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     7681 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     7741 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     7801 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     7861 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     7921 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     7981 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     8041 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     8101 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     8161 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     8221 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     8281 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     8341 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     8401 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     8461 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     8521 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     8581 gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa
     8641 cctataaaaa taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg
     8701 aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg
     8761 ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta
     8821 actatgcggc atcagagcag attgtactga gagtgcacca tatgcggtgt gaaataccgc
     8881 acagatgcgt aaggagaaaa taccgcatca ggcgccattc gccattcagg ctgcgcaact
     8941 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
     9001 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
     9061 cgacggccag tgccaagctg
//
