LOCUS       .                       8985 bp    DNA     circular UNK 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   
VERSION     
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     enhancer        54..433
                     /label="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        434..637
                     /label="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     promoter        679..697
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             747..767
                     /label="SV40 NLS"
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
     CDS             765..4868
                     /label="Cas9(H840A)"
                     /note="nickase mutant of the Cas9 endonuclease from the
                     Streptococcus pyogenes Type II CRISPR/Cas system"
     CDS             4926..4946
                     /label="SV40 NLS"
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
     CDS             6237..6257
                     /label="SV40 NLS"
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
     CDS             6267..6293
                     /label="c-myc NLS"
                     /note="nuclear localization signal of human c-Myc
                     proto-oncogene (Dang and Lee, 1988)"
     CDS             6302..6319
                     /label="6xHis"
                     /note="6xHis affinity tag"
     polyA_signal    6348..6572
                     /label="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6643..6659)
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     protein_bind    complement(6667..6683)
                     /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)."
     promoter        complement(6691..6721)
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(6736..6757)
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(7045..7633)
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(7807..8664)
                     /label="AmpR"
                     /note="β-lactamase"
     promoter        complement(8665..8769)
                     /label="AmpR promoter"
ORIGIN
        1 tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gttgacattg
       61 attattgact agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
      121 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
      181 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
      241 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
      301 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
      361 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
      421 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
      481 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
      541 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
      601 taggcgtgta cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatccg
      661 ctagagatcc gcggccgcta atacgactca ctatagggag agccgccacc atgaaacgga
      721 cagccgacgg aagcgagttc gagtcaccaa agaagaagcg gaaagtcgac aagaagtaca
      781 gcatcggcct ggacatcggc accaactctg tgggctgggc cgtgatcacc gacgagtaca
      841 aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc atcaagaaga
      901 acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc cggctgaaga
      961 gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg caagagatct
     1021 tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa gagtccttcc
     1081 tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc gtggacgagg
     1141 tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg gtggacagca
     1201 ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc aagttccggg
     1261 gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac aagctgttca
     1321 tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac gccagcggcg
     1381 tggacgccaa ggccatcctg tctgccagac tgagcaagag cagaaagctg gaaaatctga
     1441 tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt gccctgagcc
     1501 tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc aaactgcagc
     1561 tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc ggcgaccagt
     1621 acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg agcgacatcc
     1681 tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc aagagatacg
     1741 acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag ctgcctgaga
     1801 agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac attgacggcg
     1861 gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag atggacggca
     1921 ccgaggaact gctcgtgaag ctgaagagag aggacctgct gcggaagcag cggaccttcg
     1981 acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt ctgcggcggc
     2041 aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag atcctgacct
     2101 tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc gcctggatga
     2161 ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg gacaagggcg
     2221 cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg cccaacgaga
     2281 aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac gagctgacca
     2341 aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc gagcagaaaa
     2401 aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag cagctgaaag
     2461 aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc gtggaagatc
     2521 ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag gacaaggact
     2581 tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc ctgacactgt
     2641 ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg ttcgacgaca
     2701 aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg agccggaagc
     2761 tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc ctgaagtccg
     2821 acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg acctttaaag
     2881 aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag cacattgcca
     2941 atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag gtggtggacg
     3001 agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa atggccagag
     3061 agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag cggatcgaag
     3121 agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa aacacccagc
     3181 tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg tacgtggacc
     3241 aggaactgga catcaaccgg ctgtccgact acgatgtgga cgctatcgtg cctcagagct
     3301 ttctgaagga cgactccatc gacaacaagg tgctgaccag aagcgacaag aaccggggca
     3361 agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac tggcggcagc
     3421 tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag gccgagagag
     3481 gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg gaaacccggc
     3541 agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag tacgacgaga
     3601 atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg gtgtccgatt
     3661 tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac cacgcccacg
     3721 acgcctacct gaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct aagctggaaa
     3781 gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc gccaagagcg
     3841 agcaggaaat cggcaaggct accgccaagt acttcttcta cagcaacatc atgaactttt
     3901 tcaagaccga gattaccctg gccaacggcg agatccggaa gcggcctctg atcgagacaa
     3961 acggcgaaac cggggagatc gtgtgggata agggccggga ttttgccacc gtgcggaaag
     4021 tgctgagcat gccccaagtg aatatcgtga aaaagaccga ggtgcagaca ggcggcttca
     4081 gcaaagagtc tatcctgccc aagaggaaca gcgataagct gatcgccaga aagaaggact
     4141 gggaccctaa gaagtacggc ggcttcgaca gccccaccgt ggcctattct gtgctggtgg
     4201 tggccaaagt ggaaaagggc aagtccaaga aactgaagag tgtgaaagag ctgctgggga
     4261 tcaccatcat ggaaagaagc agcttcgaga agaatcccat cgactttctg gaagccaagg
     4321 gctacaaaga agtgaaaaag gacctgatca tcaagctgcc taagtactcc ctgttcgagc
     4381 tggaaaacgg ccggaagaga atgctggcct ctgccggcga actgcagaag ggaaacgaac
     4441 tggccctgcc ctccaaatat gtgaacttcc tgtacctggc cagccactat gagaagctga
     4501 agggctcccc cgaggataat gagcagaaac agctgtttgt ggaacagcac aagcactacc
     4561 tggacgagat catcgagcag atcagcgagt tctccaagag agtgatcctg gccgacgcta
     4621 atctggacaa agtgctgtcc gcctacaaca agcaccggga taagcccatc agagagcagg
     4681 ccgagaatat catccacctg tttaccctga ccaatctggg agcccctgcc gccttcaagt
     4741 actttgacac caccatcgac cggaagaggt acaccagcac caaagaggtg ctggacgcca
     4801 ccctgatcca ccagagcatc accggcctgt acgagacacg gatcgacctg tctcagctgg
     4861 gaggtgactc cggcggaagc tctggtggca gcaagcggac cgccgacggc tctgaattcg
     4921 agagccctaa gaagaaaaga aaggtgagcg gaggctctag cggcggaagc ggaagacctt
     4981 acgttacact gaatctgaac ggcatgttca tggacaagtt caagccatac agcaaatcta
     5041 atgcccctat taccaccctg gagaagctga gcaaagctct gtccatcagc gtagaggagc
     5101 tgaaggctat cgccgaactg agcctggacg agaaatacac actgaagaaa atccccaaga
     5161 tcgacggcag caagaggatt gtgtactccc tccatcctaa gatgcggctg ctgcaaagca
     5221 gaatcaacga aagaatcttc aaggaactgg tggtgttccc atcttttctg ttcggctccg
     5281 tgcctagcaa aaatgacgtg ctgaacagca atgtgaagcg ggattacgtg tcctgcgcca
     5341 aggcccactg cggcgccaaa accgtactga aagtggacat cagcaacttc ttcgacaaca
     5401 tccatagaga tctggtcaga tctgtgttcg aggaaatcct gcacatcaaa gatgaggccc
     5461 tggattacct ggtcgatatc tgcacaaagg acgacttcgt ggtccaggga gccctgacaa
     5521 gctcttatat cgccaccctg tgcctgtttg ccgtggaagg cgacgtggtg cggcgggccc
     5581 agagaaaggg actggtgtac acaagactgg tggatgacat caccgtgtct tctaaaatct
     5641 ccaactacga cttcagccag atgcagagcc acatcgagcg gatgctgtct gagcacaacc
     5701 tgcccatcaa caagcacaag accaagatct tccactgtag cagcgagcct atcaaggtgc
     5761 acggcctgat agttgactac gacagcccca gactgccttc tgataaggtg aagcgcatcc
     5821 gggccagcat ccacaacttg aagctgctgg ccgccaagaa caacaccaag accagcgtgg
     5881 cctacagaaa ggaattcaac agatgcatgg gcagagtgaa cgagctgggc cgggtgggcc
     5941 acgagaagta cgagagcttt aaaaagcagc tgcaggccat caagcctatg cctagcaatc
     6001 gggacgtggc cgtgatcgac gccgctatca agagccttga gctgagctac agcaagggca
     6061 atcagaacaa gcactggtac aagaggaagt atgatctgac cagatacaag atgatcatct
     6121 tgaccagaag cgagagcttc aaagagaagc tcgaatgttt taagtcccgg ctggctagcc
     6181 tgaagcccct gtctggcggc tcaaaaagaa ccgccgacgg cagcgaattc gagtctccca
     6241 agaagaagag gaaagtcggc tctggccctg ccgctaagag agtgaagctg gactaaccgg
     6301 tcatcatcac catcaccatt gagtttaaac ccgctgatca gcctcgactg tgccttctag
     6361 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     6421 tcccactgtc ctttcctaat aaaatgagaa aattgcatcg cattgtctga gtaggtgtca
     6481 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     6541 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     6601 ctcgataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     6661 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     6721 agcctagggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     6781 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     6841 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     6901 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     6961 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     7021 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     7081 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     7141 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     7201 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     7261 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     7321 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     7381 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     7441 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     7501 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     7561 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     7621 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacactc agtggaacga
     7681 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     7741 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     7801 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     7861 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     7921 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     7981 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     8041 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     8101 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     8161 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     8221 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     8281 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     8341 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     8401 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     8461 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     8521 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     8581 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     8641 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     8701 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     8761 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatcgatc
     8821 tcccgatccc ctagggtcga ctctcagtac aatctgctct gatgccgcat agttaagcca
     8881 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc
     8941 tacaacaagg caaggcttga ccgacaattg catgaagaat ctgct
//
