LOCUS       .                       9279 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             6531..6551
                     /label="SV40 NLS"
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
     CDS             6561..6587
                     /label="c-myc NLS"
                     /note="nuclear localization signal of human c-Myc
                     proto-oncogene (Dang and Lee, 1988)"
     CDS             6596..6613
                     /label="6xHis"
                     /note="6xHis affinity tag"
     polyA_signal    6642..6866
                     /label="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6937..6953)
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     protein_bind    complement(6961..6977)
                     /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(6985..7015)
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(7030..7051)
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(7339..7927)
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(8101..8958)
                     /label="AmpR"
                     /note="β-lactamase"
     promoter        complement(8959..9063)
                     /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 atcagcagct
     4981 ctaagcacac cctgagccag atgaacaagg tgagcaacat cgtgaaggaa cccgagctgc
     5041 ccgacatcta caaggaattt aaggacatca ccgccgacac caataccgag aagctgccta
     5101 agcctattaa gggcctggaa tttgaagtgg aactgacaca ggagaactac agactgccta
     5161 tccggaacta tcctctgact ccagtcaaga tgcaggccat gaacgacgag atcaatcaag
     5221 gcctgaaagg cggcatcatc agagagagca aggccatcaa cgcctgccct gttatattcg
     5281 tgcccagaaa ggaaggcaca ctgcgcatgg tggtcgacta caggcccctg aacaagtacg
     5341 tgaagcccaa cgtctacccc ctgccactga ttgaacaact gctggccaag atccagggca
     5401 gcaccatttt caccaagctg gacctgaaaa gcgcctacca ccagatcaga gtgcgaaagg
     5461 gagatgagca caagctggcc ttccggtgcc ccagaggagt cttcgagtac ctggtgatgc
     5521 cttacggcat cagcacagcc cctgcccact ttcagtactt catcaacaca atcctgggcg
     5581 aggccaagga aagccacgtg gtgtgctaca tggacgacat cctgatccat tccaagtccg
     5641 agtccgaaca cgtgaaacat gtgaaggacg tgctgcaaaa gctgaagaac gctaatctga
     5701 tcatcaacca ggccaaatgc gagtttcacc agagccaagt gaagttcatc ggctaccaca
     5761 tcagcgagaa gggcttaacc ccatgtcagg agaacatcga caaggtgctg cagtggaaac
     5821 agcctaaaaa ccggaaggaa ctgagacagt tcctgggcag cgtgaactac ctgagaaaat
     5881 tcatccccaa aaccagccag ttgacccacc ctctgaacaa actcctgaaa aaggatgtca
     5941 gatggaaatg gacccctacc cagacccagg ctatcgagaa tatcaagcaa tgtctggtgt
     6001 ctcctcctgt gctgaggcac ttcgacttca gcaagaagat cctgcttgag acagacgttt
     6061 ctgatgtggc cgtgggagcc gtgctgagcc agaagcatga tgatgataag tactaccctg
     6121 tgggctatta cagcgctaaa atgagcaaag cccagctgaa ttattctgtg tccgacaagg
     6181 agatgctggc tatcatcaaa agcctggagc actggcggca ctacctggaa tctacaatcg
     6241 agcccttcaa gatcctgacc gaccacagaa acctgattgg aagaatcaca aacgagagcg
     6301 aaccagagaa caagcggctg gccagatggc agctgttcct gcaggacttc aacttcgaga
     6361 tcaactacag acctggctct gcaaatcaca tcgccgatgc cctgtctaga atcgtggacg
     6421 agactgagcc tatccctaag gacaacgaag ataacagcat caacttcgtg aaccagatca
     6481 gcatctctgg cggctcaaaa agaaccgccg acggcagcga attcgagtct cccaagaaga
     6541 agaggaaagt cggctctggc cctgccgcta agagagtgaa gctggactaa ccggtcatca
     6601 tcaccatcac cattgagttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
     6661 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     6721 tgtcctttcc taataaaatg agaaaattgc atcgcattgt ctgagtaggt gtcattctat
     6781 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     6841 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctcgat
     6901 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     6961 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagccta
     7021 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     7081 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     7141 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     7201 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     7261 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     7321 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     7381 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     7441 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     7501 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     7561 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     7621 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     7681 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     7741 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     7801 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     7861 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     7921 atctcaagaa gatcctttga tcttttctac ggggtctgac actcagtgga acgaaaactc
     7981 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     8041 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     8101 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     8161 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     8221 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     8281 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     8341 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     8401 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     8461 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     8521 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     8581 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     8641 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     8701 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     8761 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     8821 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     8881 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     8941 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     9001 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     9061 gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc gatctcccga
     9121 tcccctaggg tcgactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct
     9181 gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac
     9241 aaggcaaggc ttgaccgaca attgcatgaa gaatctgct
//
