LOCUS       V022927                 8894 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V022927
VERSION     V022927
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..2937
                     /label="TIMP1(NM_003254)"
                     /note="TIMP1(NM_003254)"
                     /gene="TIMP1"
     CDS             2944..3009
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3053..3264
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3277..3545
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      3574..3583
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             3604..4266
                     /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             4336..4389
                     /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             4390..4989
                     /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    4990..5577
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5651..5884
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    5956..6077
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6096..6231
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6269..6285)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6293..6309
                     /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(6317..6347)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6362..6383
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6671..7259)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7430..8290)
                     /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(8291..8395)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     8869..8885
                     /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 gccaccatgg ccccctttga gcccctggct
     2341 tctggcatcc tgttgttgct gtggctgata gcccccagca gggcctgcac ctgtgtccca
     2401 ccccacccac agacggcctt ctgcaattcc gacctcgtca tcagggccaa gttcgtgggg
     2461 acaccagaag tcaaccagac caccttatac cagcgttatg agatcaagat gaccaagatg
     2521 tataaagggt tccaagcctt aggggatgcc gctgacatcc ggttcgtcta cacccccgcc
     2581 atggagagtg tctgcggata cttccacagg tcccacaacc gcagcgagga gtttctcatt
     2641 gctggaaaac tgcaggatgg actcttgcac atcactacct gcagttttgt ggctccctgg
     2701 aacagcctga gcttagctca gcgccggggc ttcaccaaga cctacactgt tggctgtgag
     2761 gaatgcacag tgtttccctg tttatccatc ccctgcaaac tgcagagtgg cactcattgc
     2821 ttgtggacgg accagctcct ccaaggctct gaaaagggct tccagtcccg tcaccttgcc
     2881 tgcctgcctc gggagccagg gctgtgcacc tggcagtccc tgcggtccca gatagccgga
     2941 tccgactaca aagaccatga cggtgattat aaagatcatg acatcgacta caaggatgac
     3001 gatgacaagt agcggccgcg aaggatctgc gatcgctccg gtgcccgtca gtgggcagag
     3061 cgcacatcgc ccacagtccc cgagaagttg gggggagggg tcggcaattg aacgggtgcc
     3121 tagagaaggt ggcgcggggt aaactgggaa agtgatgtcg tgtactggct ccgccttttt
     3181 cccgagggtg ggggagaacc gtatataagt gcagtagtcg ccgtgaacgt tctttttcgc
     3241 aacgggtttg ccgccagaac acagctgaag cttcgagggg ctcgcatctc tccttcacgc
     3301 gcccgccgcc ctacctgagg ccgccatcca cgccggttga gtcgcgttct gccgcctccc
     3361 gcctgtggtg cctcctgaac tgcgtccgcc gtctaggtaa gtttaaagct caggtcgaga
     3421 ccgggccttt gtccggcgct cccttggagc ctacctagac tcagccggct ctccacgctt
     3481 tgcctgaccc tgcttgctca actctacgtc tttgtttcgt tttctgttct gcgccgttac
     3541 agatccaagc tgtgaccggc gcctacgcta gacgccacca tggagagcga cgagagcggc
     3601 ctgcccgcca tggagatcga gtgccgcatc accggcaccc tgaacggcgt ggagttcgag
     3661 ctggtgggcg gcggagaggg cacccccaag cagggccgca tgaccaacaa gatgaagagc
     3721 accaaaggcg ccctgacctt cagcccctac ctgctgagcc acgtgatggg ctacggcttc
     3781 taccacttcg gcacctaccc cagcggctac gagaacccct tcctgcacgc catcaacaac
     3841 ggcggctaca ccaacacccg catcgagaag tacgaggacg gcggcgtgct gcacgtgagc
     3901 ttcagctacc gctacgaggc cggccgcgtg atcggcgact tcaaggtggt gggcaccggc
     3961 ttccccgagg acagcgtgat cttcaccgac aagatcatcc gcagcaacgc caccgtggag
     4021 cacctgcacc ccatgggcga taacgtgctg gtgggcagct tcgcccgcac cttcagcctg
     4081 cgcgacggcg gctactacag cttcgtggtg gacagccaca tgcacttcaa gagcgccatc
     4141 caccccagca tcctgcagaa cgggggcccc atgttcgcct tccgccgcgt ggaggagctg
     4201 cacagcaaca ccgagctggg catcgtggag taccagcacg ccttcaagac ccccatcgcc
     4261 ttcgccagat cccgcgctca gtcgtccaat tctgccgtgg acggcaccgc cggacccggc
     4321 tccaccggat ctcgcgaggg cagaggaagt cttctaacat gcggtgacgt ggaggagaat
     4381 cccggcccta tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc
     4441 agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc
     4501 gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc
     4561 gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc
     4621 acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag
     4681 ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg
     4741 cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag
     4801 ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc
     4861 gccttcctgg agacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc
     4921 gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc
     4981 ggtgcctgaa atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac
     5041 tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt
     5101 gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat
     5161 gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca
     5221 acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc
     5281 cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg
     5341 gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaagctgac gtcctttcca
     5401 tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct
     5461 tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt
     5521 ccgcgtctcc gccttcgccc tcagacgagt cggatctccc tttggccgcc tccccgcctg
     5581 gtacctttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa
     5641 aaggggggac tggaagggct aattcactcc caacgaaaat aagatctgct ttttgcttgt
     5701 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
     5761 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
     5821 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
     5881 agcagtagta gttcatgtca tcttattatt cagtatttat aacttgcaaa gaaatgaata
     5941 tcagagagtg agaggaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     6001 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     6061 ctcatcaatg tatcttatca tgtctggctc tagctatccc gcccctaact ccgcccagtt
     6121 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     6181 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctagactttt
     6241 gcagagacgg cccaaattcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     6301 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     6361 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     6421 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     6481 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     6541 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     6601 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     6661 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     6721 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     6781 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     6841 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     6901 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     6961 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     7021 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     7081 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     7141 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     7201 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     7261 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     7321 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     7381 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     7441 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     7501 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     7561 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     7621 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     7681 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     7741 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     7801 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     7861 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     7921 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     7981 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     8041 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     8101 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     8161 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     8221 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     8281 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     8341 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     8401 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     8461 aaaataggcg tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc
     8521 tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca
     8581 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg
     8641 cggcatcaga gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat
     8701 gcgtaaggag aaaataccgc atcaggcgcc attcgccatt caggctgcgc aactgttggg
     8761 aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg
     8821 caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg
     8881 ccagtgccaa gctg
//
