LOCUS       .                      11128 bp    DNA     circular UNK 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   
VERSION     
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     CDS             1..21
                     /label="SV40 NLS"
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
     CDS             19..4122
                     /label="Cas9(H840A)"
                     /note="nickase mutant of the Cas9 endonuclease from the
                     Streptococcus pyogenes Type II CRISPR/Cas system"
     CDS             4180..4200
                     /label="SV40 NLS"
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
     CDS             6301..6321
                     /label="SV40 NLS"
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
     CDS             6331..6357
                     /label="c-myc NLS"
                     /note="nuclear localization signal of human c-Myc
                     proto-oncogene (Dang and Lee, 1988)"
     CDS             6424..7140
                     /label="EGFP"
                     /note="the original enhanced GFP (Yang et al., 1996)"
     CDS             7149..7166
                     /label="6xHis"
                     /note="6xHis affinity tag"
     polyA_signal    7195..7419
                     /label="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     promoter        complement(7538..7568)
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(7583..7604)
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(7892..8480)
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(8654..9511)
                     /label="AmpR"
                     /note="β-lactamase"
     promoter        complement(9512..9616)
                     /label="AmpR promoter"
     intron          10111..11049
                     /label="EF-1α intron A"
                     /note="intron upstream of the start codon of human EF-1α"
     promoter        11061..11079
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ccaaagaaga agcggaaagt cgacaagaag tacagcatcg gcctggacat cggcaccaac
       61 tctgtgggct gggccgtgat caccgacgag tacaaggtgc ccagcaagaa attcaaggtg
      121 ctgggcaaca ccgaccggca cagcatcaag aagaacctga tcggagccct gctgttcgac
      181 agcggcgaaa cagccgaggc cacccggctg aagagaaccg ccagaagaag atacaccaga
      241 cggaagaacc ggatctgcta tctgcaagag atcttcagca acgagatggc caaggtggac
      301 gacagcttct tccacagact ggaagagtcc ttcctggtgg aagaggataa gaagcacgag
      361 cggcacccca tcttcggcaa catcgtggac gaggtggcct accacgagaa gtaccccacc
      421 atctaccacc tgagaaagaa actggtggac agcaccgaca aggccgacct gcggctgatc
      481 tatctggccc tggcccacat gatcaagttc cggggccact tcctgatcga gggcgacctg
      541 aaccccgaca acagcgacgt ggacaagctg ttcatccagc tggtgcagac ctacaaccag
      601 ctgttcgagg aaaaccccat caacgccagc ggcgtggacg ccaaggccat cctgtctgcc
      661 agactgagca agagcagaaa gctggaaaat ctgatcgccc agctgcccgg cgagaagaag
      721 aatggcctgt tcggaaacct gattgccctg agcctgggcc tgacccccaa cttcaagagc
      781 aacttcgacc tggccgagga tgccaaactg cagctgagca aggacaccta cgacgacgac
      841 ctggacaacc tgctggccca gatcggcgac cagtacgccg acctgtttct ggccgccaag
      901 aacctgtccg acgccatcct gctgagcgac atcctgagag tgaacaccga gatcaccaag
      961 gcccccctga gcgcctctat gatcaagaga tacgacgagc accaccagga cctgaccctg
     1021 ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca aagagatttt cttcgaccag
     1081 agcaagaacg gctacgccgg ctacattgac ggcggagcca gccaggaaga gttctacaag
     1141 ttcatcaagc ccatcctgga aaagatggac ggcaccgagg aactgctcgt gaagctgaag
     1201 agagaggacc tgctgcggaa gcagcggacc ttcgacaacg gcagcatccc ccaccagatc
     1261 cacctgggag agctgcacgc cattctgcgg cggcaggaag atttttaccc attcctgaag
     1321 gacaaccggg aaaagatcga gaagatcctg accttccgca tcccctacta cgtgggccct
     1381 ctggccaggg gaaacagcag attcgcctgg atgaccagaa agagcgagga aaccatcacc
     1441 ccctggaact tcgaggaagt ggtggacaag ggcgcttccg cccagagctt catcgagcgg
     1501 atgaccaact tcgataagaa cctgcccaac gagaaggtgc tgcccaagca cagcctgctg
     1561 tacgagtact tcaccgtgta taacgagctg accaaagtga aatacgtgac cgagggaatg
     1621 agaaagcccg ccttcctgag cggcgagcag aaaaaggcca tcgtggacct gctgttcaag
     1681 accaaccgga aagtgaccgt gaagcagctg aaagaggact acttcaagaa aatcgagtgc
     1741 ttcgactccg tggaaatctc cggcgtggaa gatcggttca acgcctccct gggcacatac
     1801 cacgatctgc tgaaaattat caaggacaag gacttcctgg acaatgagga aaacgaggac
     1861 attctggaag atatcgtgct gaccctgaca ctgtttgagg acagagagat gatcgaggaa
     1921 cggctgaaaa cctatgccca cctgttcgac gacaaagtga tgaagcagct gaagcggcgg
     1981 agatacaccg gctggggcag gctgagccgg aagctgatca acggcatccg ggacaagcag
     2041 tccggcaaga caatcctgga tttcctgaag tccgacggct tcgccaacag aaacttcatg
     2101 cagctgatcc acgacgacag cctgaccttt aaagaggaca tccagaaagc ccaggtgtcc
     2161 ggccagggcg atagcctgca cgagcacatt gccaatctgg ccggcagccc cgccattaag
     2221 aagggcatcc tgcagacagt gaaggtggtg gacgagctcg tgaaagtgat gggccggcac
     2281 aagcccgaga acatcgtgat cgaaatggcc agagagaacc agaccaccca gaagggacag
     2341 aagaacagcc gcgagagaat gaagcggatc gaagagggca tcaaagagct gggcagccag
     2401 atcctgaaag aacaccccgt ggaaaacacc cagctgcaga acgagaagct gtacctgtac
     2461 tacctgcaga atgggcggga tatgtacgtg gaccaggaac tggacatcaa ccggctgtcc
     2521 gactacgatg tggacgctat cgtgcctcag agctttctga aggacgactc catcgacaac
     2581 aaggtgctga ccagaagcga caagaaccgg ggcaagagcg acaacgtgcc ctccgaagag
     2641 gtcgtgaaga agatgaagaa ctactggcgg cagctgctga acgccaagct gattacccag
     2701 agaaagttcg acaatctgac caaggccgag agaggcggcc tgagcgaact ggataaggcc
     2761 ggcttcatca agagacagct ggtggaaacc cggcagatca caaagcacgt ggcacagatc
     2821 ctggactccc ggatgaacac taagtacgac gagaatgaca agctgatccg ggaagtgaaa
     2881 gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga aggatttcca gttttacaaa
     2941 gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct acctgaacgc cgtcgtggga
     3001 accgccctga tcaaaaagta ccctaagctg gaaagcgagt tcgtgtacgg cgactacaag
     3061 gtgtacgacg tgcggaagat gatcgccaag agcgagcagg aaatcggcaa ggctaccgcc
     3121 aagtacttct tctacagcaa catcatgaac tttttcaaga ccgagattac cctggccaac
     3181 ggcgagatcc ggaagcggcc tctgatcgag acaaacggcg aaaccgggga gatcgtgtgg
     3241 gataagggcc gggattttgc caccgtgcgg aaagtgctga gcatgcccca agtgaatatc
     3301 gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag agtctatcct gcccaagagg
     3361 aacagcgata agctgatcgc cagaaagaag gactgggacc ctaagaagta cggcggcttc
     3421 gacagcccca ccgtggccta ttctgtgctg gtggtggcca aagtggaaaa gggcaagtcc
     3481 aagaaactga agagtgtgaa agagctgctg gggatcacca tcatggaaag aagcagcttc
     3541 gagaagaatc ccatcgactt tctggaagcc aagggctaca aagaagtgaa aaaggacctg
     3601 atcatcaagc tgcctaagta ctccctgttc gagctggaaa acggccggaa gagaatgctg
     3661 gcctctgccg gcgaactgca gaagggaaac gaactggccc tgccctccaa atatgtgaac
     3721 ttcctgtacc tggccagcca ctatgagaag ctgaagggct cccccgagga taatgagcag
     3781 aaacagctgt ttgtggaaca gcacaagcac tacctggacg agatcatcga gcagatcagc
     3841 gagttctcca agagagtgat cctggccgac gctaatctgg acaaagtgct gtccgcctac
     3901 aacaagcacc gggataagcc catcagagag caggccgaga atatcatcca cctgtttacc
     3961 ctgaccaatc tgggagcccc tgccgccttc aagtactttg acaccaccat cgaccggaag
     4021 aggtacacca gcaccaaaga ggtgctggac gccaccctga tccaccagag catcaccggc
     4081 ctgtacgaga cacggatcga cctgtctcag ctgggaggtg actccggcgg aagctctggt
     4141 ggcagcaagc ggaccgccga cggctctgaa ttcgagagcc ctaagaagaa aagaaaggtg
     4201 agcggaggct ctagcggcgg aagcaccctg aacattgaag acgagtatag actgcatgaa
     4261 acaagcaagg aacccgacgt gtccctgggc tccacctggc tgtccgactt tccccaggcc
     4321 tgggccgaga caggaggaat gggcctggcc gtgcggcagg cacccctgat catccctctg
     4381 aaggccacct ctacacccgt gagcatcaag cagtacccta tgtctcagga ggccagactg
     4441 ggcatcaagc ctcacatcca gaggctgctg gaccagggca tcctggtgcc atgccagagc
     4501 ccctggaaca caccactgct gcccgtgaag aagccaggca ccaatgacta tagacccgtg
     4561 caggatctga gagaggtgaa caagagggtg gaggatatcc accccaccgt gcccaaccct
     4621 tacaatctgc tgtccggcct gcccccttct caccagtggt atacagtgct ggacctgaag
     4681 gatgccttct tttgtctgag actgcaccct accagccagc cactgttcgc ctttgagtgg
     4741 agggaccctg agatgggcat ctctggccag ctgacctgga cacgcctgcc tcagggcttc
     4801 aagaatagcc caacactgtt taacgaggcc ctgcaccgcg acctggcaga tttccggatc
     4861 cagcacccag atctgatcct gctgcagtac gtggacgatc tgctgctggc cgccaccagc
     4921 gagctggatt gccagcaggg aacacgcgcc ctgctgcaga ccctgggaaa cctgggatat
     4981 agggcatccg ccaagaaggc ccagatctgt cagaagcagg tgaagtacct gggctatctg
     5041 ctgaaggagg gccagagatg gctgacagag gccaggaagg agacagtgat gggccagcca
     5101 acacccaaga ccccaagaca gctgagggag ttcctgggca aagcaggatt ttgcaggctg
     5161 ttcatcccag gattcgcaga gatggcagca cctctgtacc cactgaccaa gccgggcacc
     5221 ctgtttaatt ggggccctga ccagcagaag gcctatcagg agatcaagca ggccctgctg
     5281 acagcaccag ccctgggcct gccagacctg accaagcctt tcgagctgtt tgtggatgag
     5341 aagcagggct acgccaaggg cgtgctgacc cagaagctgg gaccatggag acggcccgtg
     5401 gcctatctgt ccaagaagct ggacccagtg gcagcaggat ggccaccatg cctgaggatg
     5461 gtggcagcaa tcgccgtgct gacaaaggat gccggcaagc tgaccatggg acagccactg
     5521 gtcatcctgg caccacacgc agtggaggcc ctggtgaagc agcctccaga tcgctggctg
     5581 tctaacgccc ggatgacaca ctaccaggcc ctgctgctgg acaccgatcg cgtgcagttt
     5641 ggccctgtgg tggccctgaa tccagccacc ctgctgcctc tgccagagga gggcctgcag
     5701 cacaactgtc tggacatcct ggcagaggca cacggaacaa ggccagacct gaccgatcag
     5761 cccctgcctg acgccgatca cacatggtat accgatggaa gctccctgct gcaggagggc
     5821 cagaggaagg caggagcagc agtgaccaca gagacagaag tgatctgggc caaggccctg
     5881 ccagcaggca catccgccca gcgggccgag ctgatcgccc tgacccaggc cctgaagatg
     5941 gccgagggca agaagctgaa cgtgtacaca gactccagat atgccttcgc caccgcacac
     6001 atccacggag agatctacag gcgccggggc tggctgacct ctgagggcaa ggagatcaag
     6061 aacaaggatg agatcctggc cctgctgaag gccctgtttc tgcccaagcg gctgagcatc
     6121 atccactgtc ctggacacca gaagggacac tccgccgagg caaggggcaa tcggatggcc
     6181 gaccaggccg ccagaaaggc tgctattact gaaactcccg acacttccac tctgctgatt
     6241 gaaaactcct ccccttctgg cggctcaaaa agaaccgccg acggcagcga attcgagtct
     6301 cccaagaaga agaggaaagt cggctctggc cctgccgcta agagagtgaa gctggacgga
     6361 tccggcgcaa caaacttctc tctgctgaaa caagccggag atgtcgaaga gaatcctgga
     6421 ccgatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
     6481 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
     6541 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
     6601 accctcgtga ccaccctgac ctatggagtg cagtgcttca gccgctaccc cgaccacatg
     6661 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
     6721 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
     6781 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     6841 cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga caagcagaag
     6901 aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc
     6961 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     7021 cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg
     7081 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
     7141 taaccggtca tcatcaccat caccattgag tttaaacccg ctgatcagcc tcgactgtgc
     7201 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag
     7261 gtgccactcc cactgtcctt tcctaataaa atgagaaaat tgcatcgcat tgtctgagta
     7321 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag
     7381 acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca
     7441 gctggggctc gataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
     7501 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     7561 agtgtaaagc ctagggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     7621 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     7681 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     7741 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     7801 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     7861 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     7921 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     7981 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     8041 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     8101 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     8161 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     8221 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     8281 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     8341 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     8401 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     8461 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacactcagt
     8521 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     8581 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     8641 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     8701 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     8761 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     8821 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     8881 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     8941 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     9001 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     9061 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     9121 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     9181 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     9241 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     9301 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     9361 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     9421 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     9481 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     9541 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     9601 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag
     9661 atcgatctcc cgatccccta gggtcgactc tcagtacaat ctgctctgat gccgcatagt
     9721 taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa
     9781 tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg cttagggtta
     9841 ggcgttttgc gctgcttcgc gatgtacggg ccagatatag gctccggtgc ccgtcagtgg
     9901 gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg caattgaacc
     9961 ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc
    10021 ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt
    10081 tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc ccgcgggcct
    10141 ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacctg gctgcagtac
    10201 gtgattcttg atcccgagct tcgggttgga agtgggtggg agagttcgag gccttgcgct
    10261 taaggagccc cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg gggccgccgc
    10321 gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc tctagccatt
    10381 taaaattttt gatgacctgc tgcgacgctt tttttctggc aagatagtct tgtaaatgcg
    10441 ggccaagatc tgcacactgg tatttcggtt tttggggccg cgggcggcga cggggcccgt
    10501 gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc gagaatcgga
    10561 cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc gccgtgtatc
    10621 gccccgccct gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc ggaaagatgg
    10681 ccgcttcccg gccctgctgc agggagctca aaatggagga cgcggcgctc gggagagcgg
    10741 gcgggtgagt cacccacaca aaggaaaagg gcctttccgt cctcagccgt cgcttcatgt
    10801 gactccacgg agtaccgggc gccgtccagg cacctcgatt agttctcgag cttttggagt
    10861 acgtcgtctt taggttgggg ggaggggttt tatgcgatgg agtttcccca cactgagtgg
    10921 gtggagactg aagttaggcc agcttggcac ttgatgtaat tctccttgga atttgccctt
    10981 tttgagtttg gatcttggtt cattctcaag cctcagacag tggttcaaag tttttttctt
    11041 ccatttcagg tgtcgtgagc taatacgact cactataggg agagccgcca ccatgaaacg
    11101 gacagccgac ggaagcgagt tcgagtca
//
