LOCUS       V025603                 9947 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V025603
VERSION     V025603
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     LTR             1..634
                     /note="3' long terminal repeat (LTR) from HIV-1"
                     /label="3' LTR"
     misc_feature    681..806
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
                     /label="HIV-1 Ψ"
     misc_feature    1303..1536
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
                     /label="RRE"
     CDS             1721..1765
                     /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    2028..2143
                     /note="central polypurine tract and central termination
                     sequence of HIV-1 (lacking the first T)"
                     /label="cPPT/CTS"
     promoter        2337..3519
                     /note="strong constitutive promoter for human elongation
                     factor EF-1α"
                     /label="EF-1α promoter"
     CDS             3551..3568
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /transl_table=1
                     /translation="HHHHHH"
                     /label="6xHis"
     CDS             3569..4606
                     /label="HCAR1(NM_032554)"
                     /note="HCAR1(NM_032554)"
                     /gene="HCAR1"
     CDS             4613..4678
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     misc_feature    4697..5283
                     /note="internal ribosome entry site (IRES) of the
                     encephalomyocarditis virus (EMCV)"
                     /label="IRES2"
     CDS             5303..5902
                     /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    5916..6504
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6711..7344
                     /note="3' long terminal repeat (LTR) from HIV-1"
                     /label="3' LTR"
     primer_bind     complement(7472..7488)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    7496..7512
                     /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(7520..7550)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    7565..7586
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7874..8462)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8633..9493)
                     /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(9494..9598)
                     /gene="bla"
                     /label="AmpR promoter"
     polyA_signal    9646..9780
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccttt aaaagaaaag
     2041 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2101 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
     2161 acagcagaga tccagtttat cgatgagtaa ttcatacaaa aggactcgcc cctgccttgg
     2221 ggaatcccag ggaccgtcgt taaactccca ctaacgtaga acccagagat cgctgcgttc
     2281 ccgccccctc acccgcccgc tctcgtcatc actgaggtgg agaagagcat gcgtgaggct
     2341 ccggtgcccg tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag
     2401 gggtcggcaa ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg
     2461 tcgtgtactg gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag
     2521 tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag aacacaggta agtgccgtgt
     2581 gtggttcccg cgggcctggc ctctttacgg gttatggccc ttgcgtgcct tgaattactt
     2641 ccacgcccct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
     2701 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
     2761 cttgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
     2821 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
     2881 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
     2941 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
     3001 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
     3061 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     3121 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     3181 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     3241 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     3301 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     3361 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     3421 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     3481 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtacgaatt
     3541 cgccaccatg caccatcacc atcaccatat gtacaacggg tcgtgctgcc gcatcgaggg
     3601 ggacaccatc tcccaggtga tgccgccgct gctcattgtg gcctttgtgc tgggcgcact
     3661 aggcaatggg gtcgccctgt gtggtttctg cttccacatg aagacctgga agcccagcac
     3721 tgtttacctt ttcaatttgg ccgtggctga tttcctcctt atgatctgcc tgccttttcg
     3781 gacagactat tacctcagac gtagacactg ggcttttggg gacattccct gccgagtggg
     3841 gctcttcacg ttggccatga acagggccgg gagcatcgtg ttccttacgg tggtggctgc
     3901 ggacaggtat ttcaaagtgg tccaccccca ccacgcggtg aacactatct ccacccgggt
     3961 ggcggctggc atcgtctgca ccctgtgggc cctggtcatc ctgggaacag tgtatctttt
     4021 gctggagaac catctctgcg tgcaagagac ggccgtctcc tgtgagagct tcatcatgga
     4081 gtcggccaat ggctggcatg acatcatgtt ccagctggag ttctttatgc ccctcggcat
     4141 catcttattt tgctccttca agattgtttg gagcctgagg cggaggcagc agctggccag
     4201 acaggctcgg atgaagaagg cgacccggtt catcatggtg gtggcaattg tgttcatcac
     4261 atgctacctg cccagcgtgt ctgctagact ctatttcctc tggacggtgc cctcgagtgc
     4321 ctgcgatccc tctgtccatg gggccctgca cataaccctc agcttcacct acatgaacag
     4381 catgctggat cccctggtgt attatttttc aagcccctcc tttcccaaat tctacaacaa
     4441 gctcaaaatc tgcagtctga aacccaagca gccaggacac tcaaaaacac aaaggccgga
     4501 agagatgcca atttcgaacc tcggtcgcag gagttgcatc agtgtggcaa atagtttcca
     4561 aagccagtct gatgggcaat gggatcccca cattgttgag tggcactcta gagactacaa
     4621 agaccatgac ggtgattata aagatcatga catcgactac aaggatgacg atgacaagta
     4681 gcggccgcgg atcccgcccc tctccctccc ccccccctaa cgttactggc cgaagccgct
     4741 tggaataagg ccggtgtgcg tttgtctata tgttattttc caccatattg ccgtcttttg
     4801 gcaatgtgag ggcccggaaa cctggccctg tcttcttgac gagcattcct aggggtcttt
     4861 cccctctcgc caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca gttcctctgg
     4921 aagcttcttg aagacaaaca acgtctgtag cgaccctttg caggcagcgg aaccccccac
     4981 ctggcgacag gtgcctctgc ggccaaaagc cacgtgtata agatacacct gcaaaggcgg
     5041 cacaacccca gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa tggctctcct
     5101 caagcgtatt caacaagggg ctgaaggatg cccagaaggt accccattgt atgggatctg
     5161 atctggggcc tcggtgcaca tgctttacat gtgtttagtc gaggttaaaa aaacgtctag
     5221 gccccccgaa ccacggggac gtggttttcc tttgaaaaac acgatgataa gcttgccaca
     5281 acccacaagg agacgacctt ccatgaccga gtacaagccc acggtgcgcc tcgccacccg
     5341 cgacgacgtc ccccgggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac
     5401 gcgccacacc gtcgacccgg accgccacat cgagcgggtc accgagctgc aagaactctt
     5461 cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt
     5521 ggcggtctgg accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc
     5581 gcgcatggcc gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct
     5641 ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga
     5701 ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg
     5761 cgccggggtg cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg
     5821 gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat
     5881 gacccgcaag cccggtgcct agacgcgtct ggaacaatca acctctggat tacaaaattt
     5941 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     6001 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     6061 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     6121 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     6181 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     6241 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     6301 tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg tgttgccacc tggattctgc
     6361 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     6421 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga
     6481 tctccctttg ggccgcctcc ccgcctggaa ttaattctgc agtcgagacc tagaaaaaca
     6541 tggagcaatc acaagtagca atacagcagc taccaatgct gattgtgcct ggctagaagc
     6601 acaagaggag gaggaggtgg gttttccagt cacacctcag gtacctttaa gaccaatgac
     6661 ttacaaggca gctgtagatc ttagccactt tttaaaagaa aagaggggac tggaagggct
     6721 aattcactcc caacgaagac aagatatcct tgatctgtgg atctaccaca cacaaggcta
     6781 cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac tgacctttgg
     6841 atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca ataaaggaga
     6901 gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg agagagaagt
     6961 gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag agctgcatcc
     7021 ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg ctggggactt
     7081 tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat cctgcatata
     7141 agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga gcctgggagc
     7201 tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc
     7261 aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt
     7321 agtcagtgtg gaaaatctct agcagtagta gttcatgtca tcttattatt cagtatttat
     7381 aacttgcaaa gaaatgaata tcagagagtg agaggccttg acattgctag cgtttaccgt
     7441 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     7501 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     7561 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     7621 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     7681 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     7741 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     7801 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     7861 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     7921 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     7981 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     8041 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     8101 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     8161 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     8221 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     8281 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     8341 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     8401 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     8461 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     8521 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     8581 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     8641 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     8701 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     8761 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     8821 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     8881 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     8941 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     9001 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     9061 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     9121 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     9181 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     9241 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     9301 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     9361 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     9421 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     9481 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     9541 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     9601 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatcaactt gtttattgca
     9661 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     9721 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc
     9781 aactggataa ctcaagctaa ccaaaatcat cccaaacttc ccaccccata ccctattacc
     9841 actgccaatt acctgtggtt tcatttactc taaacctgtg attcctctga attattttca
     9901 ttttaaagaa attgtatttg ttaaatatgt actacaaact tagtagt
//
