LOCUS       .                       8789 bp    DNA     circular UNK 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   
VERSION     
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     promoter        162..402
                     /label="U6 promoter"
                     /note="RNA polymerase III promoter for human U6 snRNA"
     promoter        546..859
                     /label="U6 promoter"
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das
                     et al., 1988)"
     misc_RNA        898..973
                     /label="gRNA scaffold"
                     /note="guide RNA scaffold for the Streptococcus pyogenes
                     CRISPR/Cas9 system"
     misc_feature    1029..1146
                     /label="cPPT/CTS"
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
     promoter        1285..2463
                     /label="EF-1α promoter"
                     /note="strong constitutive promoter for human elongation
                     factor EF-1α"
     CDS             2476..3072
                     /label="PuroR"
                     /note="puromycin N-acetyltransferase"
     misc_feature    3091..3679
                     /label="WPRE"
                     /note="woodchuck hepatitis virus posttranscriptional
                     regulatory element"
     LTR             3751..3984
                     /label="3' LTR (ΔU3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4062..4196
                     /label="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     rep_origin      4223..4358
                     /label="SV40 ori"
                     /note="SV40 origin of replication"
     promoter        complement(4379..4397)
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4407..4423)
                     /label="M13 fwd"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     rep_origin      4565..5020
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        5046..5150
                     /label="AmpR promoter"
     CDS             5151..6008
                     /label="AmpR"
                     /note="β-lactamase"
     rep_origin      6182..6770
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     protein_bind    7058..7079
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        7094..7124
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    7132..7148
                     /label="lac operator"
                     /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)."
     primer_bind     7156..7172
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     promoter        7193..7211
                     /label="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        7239..7465
                     /label="RSV promoter"
                     /note="Rous sarcoma virus enhancer/promoter"
     misc_feature    7693..7818
                     /label="HIV-1 Ψ"
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    8311..8544
                     /label="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
     CDS             8729..8773
                     /label="gp41 peptide"
                     /note="antigenic peptide corresponding to amino acids 655
                     to 669 of the HIV envelope protein gp41 (Lutje Hulsik et
                     al., 2013)"
ORIGIN
        1 taacaaattg gctgtggtat ataaaattat tcataatgat agtaggaggc ttggtaggtt
       61 taagaatagt ttttgctgta ctttctatag tgaatagagt taggcaggga tattcaccat
      121 tatcgtttca gacccacctc ccaaccccga ggggacccag agagggccta tttcccatga
      181 ttccttcata tttgcatata cgatacaagg ctgttagaga gataattaga attaatttga
      241 ctgtaaacac aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt
      301 agtttgcagt tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa
      361 agtatttcga tttcttggct ttatatatct tgtggaaagg actctagaac caggatgggc
      421 accacccgtt taagagctaa gctggaaaca gcatagcaag tttaaataag gctagtccgt
      481 tatcaacttg aaaaagtggc accgagtcgg tgcttttttt gtactgagtc gcccagtctc
      541 agatagatcc gacgccgcca tctctaggcc cgcgccggcc ccctcgcaca gacttgtggg
      601 agaagctcgg ctactcccct gccccggtta atttgcatat aatatttcct agtaactata
      661 gaggcttaat gtgcgataaa agacagataa tctgttcttt ttaatactag ctacatttta
      721 catgataggc ttggatttct ataagagata caaatactaa attattattt taaaaaacag
      781 cacaaaagga aactcaccct aactgtaaag taattgtgtg ttttgagact ataaatatcc
      841 cttggagaaa agccttgttc accggagacg gcttcttgct cttttttgta cgtctctgtt
      901 ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtggc
      961 accgagtcgg tgctttttta agcttggcgt aactagatct tgagacaaat ggcagtattc
     1021 atccacaatt ttaaaagaaa aggggggatt ggggggtaca gtgcagggga aagaatagta
     1081 gacataatag caacagacat acaaactaaa gaattacaaa aacaaattac aaaaattcaa
     1141 aattttcggg tttattacag ggacagcaga gatccacttt ggcgccggct cgagggggcc
     1201 cgggtgcaaa gatggataaa gttttaaaca gagaggaatc tttgcagcta atggaccttc
     1261 taggtcttga aaggagtggg aattggctcc ggtgcccgtc agtgggcaga gcgcacatcg
     1321 cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt gatccggtgc ctagagaagg
     1381 tggcgcgggg taaactggga aagtgatgtc gtgtactggc tccgcctttt tcccgagggt
     1441 gggggagaac cgtatataag tgcagtagtc gccgtgaacg ttctttttcg caacgggttt
     1501 gccgccagaa cacaggtaag tgccgtgtgt ggttcccgcg ggcctggcct ctttacgggt
     1561 tatggccctt gcgtgccttg aattacttcc acctggctgc agtacgtgat tcttgatccc
     1621 gagcttcggg ttggaagtgg gtgggagagt tcgaggcctt gcgcttaagg agccccttcg
     1681 cctcgtgctt gagttgaggc ctggcctggg cgctggggcc gccgcgtgcg aatctggtgg
     1741 caccttcgcg cctgtctcgc tgctttcgat aagtctctag ccatttaaaa tttttgatga
     1801 cctgctgcga cgcttttttt ctggcaagat agtcttgtaa atgcgggcca agatctgcac
     1861 actggtattt cggtttttgg ggccgcgggc ggcgacgggg cccgtgcgtc ccagcgcaca
     1921 tgttcggcga ggcggggcct gcgagcgcgg ccaccgagaa tcggacgggg gtagtctcaa
     1981 gctggccggc ctgctctggt gcctggcctc gcgccgccgt gtatcgcccc gccctgggcg
     2041 gcaaggctgg cccggtcggc accagttgcg tgagcggaaa gatggccgct tcccggccct
     2101 gctgcaggga gctcaaaatg gaggacgcgg cgctcgggag agcgggcggg tgagtcaccc
     2161 acacaaagga aaagggcctt tccgtcctca gccgtcgctt catgtgactc cacggagtac
     2221 cgggcgccgt ccaggcacct cgattagttc tcgagctttt ggagtacgtc gtctttaggt
     2281 tggggggagg ggttttatgc gatggagttt ccccacactg agtgggtgga gactgaagtt
     2341 aggccagctt ggcacttgat gtaattctcc ttggaatttg ccctttttga gtttggatct
     2401 tggttcattc tcaagcctca gacagtggtt caaagttttt ttcttccatt tcaggtgtcg
     2461 tgacgtacgg ccaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
     2521 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
     2581 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     2641 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc cgtggcggtc
     2701 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     2761 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
     2821 caccggccca aggagcccgc gtggttcctg gccaccgtcg gagtctcgcc cgaccaccag
     2881 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
     2941 gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc
     3001 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     3061 aagcccggtg cctgaacgcg ttaagtcgac aatcaacctc tggattacaa aatttgtgaa
     3121 agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta
     3181 atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa
     3241 tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg
     3301 tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc
     3361 ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc
     3421 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg
     3481 gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg
     3541 acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg
     3601 ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc
     3661 ctttgggccg cctccccgcg tcgactttaa gaccaatgac ttacaaggca gctgtagatc
     3721 ttagccactt tttaaaagaa aaggggggac tggaagggct aattcactcc caacgaagac
     3781 aagatctgct ttttgcttgt actgggtctc tctggttaga ccagatctga gcctgggagc
     3841 tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc
     3901 aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt
     3961 agtcagtgtg gaaaatctct agcagtacgt atagtagttc atgtcatctt attattcagt
     4021 atttataact tgcaaagaaa tgaatatcag agagtgagag gaacttgttt attgcagctt
     4081 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     4141 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggctctagc
     4201 tatcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     4261 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga
     4321 gctattccag aagtagtgag gaggcttttt tggaggccta gggacgtacc caattcgccc
     4381 tatagtgagt cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa
     4441 aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt
     4501 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa
     4561 tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     4621 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     4681 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     4741 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     4801 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     4861 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     4921 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     4981 tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa
     5041 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     5101 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     5161 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     5221 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     5281 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     5341 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     5401 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     5461 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     5521 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     5581 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     5641 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     5701 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     5761 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     5821 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     5881 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     5941 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     6001 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     6061 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     6121 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     6181 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     6241 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     6301 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     6361 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     6421 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     6481 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     6541 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     6601 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     6661 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     6721 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     6781 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     6841 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     6901 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa
     6961 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
     7021 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt
     7081 aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg
     7141 gataacaatt tcacacagga aacagctatg accatgatta cgccaagcgc gcaattaacc
     7201 ctcactaaag ggaacaaaag ctggagctgc aagcttaatg tagtcttatg caatactctt
     7261 gtagtcttgc aacatggtaa cgatgagtta gcaacatgcc ttacaaggag agaaaaagca
     7321 ccgtgcatgc cgattggtgg aagtaaggtg gtacgatcgt gccttattag gaaggcaaca
     7381 gacgggtctg acatggattg gacgaaccac tgaattgccg cattgcagag atattgtatt
     7441 taagtgccta gctcgataca taaacgggtc tctctggtta gaccagatct gagcctggga
     7501 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct
     7561 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt
     7621 ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg
     7681 aaaccagagg agctctctcg acgcaggact cggcttgctg aagcgcgcac ggcaagaggc
     7741 gaggggcggc gactggtgag tacgccaaaa attttgacta gcggaggcta gaaggagaga
     7801 gatgggtgcg agagcgtcag tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt
     7861 cggttaaggc cagggggaaa gaaaaaatat aaattaaaac atatagtatg ggcaagcagg
     7921 gagctagaac gattcgcagt taatcctggc ctgttagaaa catcagaagg ctgtagacaa
     7981 atactgggac agctacaacc atcccttcag acaggatcag aagaacttag atcattatat
     8041 aatacagtag caaccctcta ttgtgtgcat caaaggatag agataaaaga caccaaggaa
     8101 gctttagaca agatagagga agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc
     8161 gctgatcttc agacctggag gaggagatat gagggacaat tggagaagtg aattatataa
     8221 atataaagta gtaaaaattg aaccattagg agtagcaccc accaaggcaa agagaagagt
     8281 ggtgcagaga gaaaaaagag cagtgggaat aggagctttg ttccttgggt tcttgggagc
     8341 agcaggaagc actatgggcg cagcgtcaat gacgctgacg gtacaggcca gacaattatt
     8401 gtctggtata gtgcagcagc agaacaattt gctgagggct attgaggcgc aacagcatct
     8461 gttgcaactc acagtctggg gcatcaagca gctccaggca agaatcctgg ctgtggaaag
     8521 atacctaaag gatcaacagc tcctggggat ttggggttgc tctggaaaac tcatttgcac
     8581 cactgctgtg ccttggaatg ctagttggag taataaatct ctggaacaga tttggaatca
     8641 cacgacctgg atggagtggg acagagaaat taacaattac acaagcttaa tacactcctt
     8701 aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg aattagataa
     8761 atgggcaagt ttgtggaatt ggtttaaca
//
