LOCUS       .                      10439 bp    DNA     linear   UNK 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   
VERSION     
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     misc_RNA        337..412
                     /label="gRNA scaffold"
                     /note="guide RNA scaffold for the Streptococcus pyogenes
                     CRISPR/Cas9 system"
     promoter        503..743
                     /label="U6 promoter"
                     /note="RNA polymerase III promoter for human U6 snRNA"
     enhancer        943..1228
                     /label="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer;
                     contains an 18-bp deletion relative to the standard CMV
                     enhancer"
     promoter        1230..1507
                     /label="chicken β-actin promoter"
     CDS             1873..5973
                     /label="Cas9"
                     /note="S. pyogenes Cas9, codon optimized for C. elegans
                     with synthetic introns"
     CDS             6754..7470
                     /label="EGFP"
                     /note="the original enhanced GFP (Yang et al., 1996)"
     polyA_signal    7504..7711
                     /label="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   7720..7860
                     /label="AAV2 ITR"
                     /note="inverted terminal repeat of adeno-associated virus
                     serotype 2"
     rep_origin      7935..8390
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        8672..8776
                     /label="AmpR promoter"
     CDS             8777..9634
                     /label="AmpR"
                     /note="β-lactamase"
ORIGIN
        1 ggccttttgc tcacatgcgc cattctcgag catgcgccat tctcgagcat gctcgagaat
       61 ggcgcatgtg agggcctatt tcccatgatt ccttcatatt tgcatatacg atacaaggct
      121 gttagagaga taattggaat taatttgact gtaaacacaa agatattagt acaaaatacg
      181 tgacgtagaa agtaataatt tcttgggtag tttgcagttt taaaattatg ttttaaaatg
      241 gactatcata tgcttaccgt aacttgaaag tatttcgatt tcttggcttt atatatcttg
      301 tggaaaggac gaaacaccgg gtcttcgaga agacctgttt tagagctaga aatagcaagt
      361 taaaataagg ctagtccgtt atcaacttga aaaagtggca ccgagtcggt gcttttttgt
      421 tttagagcta gaaatagcaa gttaaaataa ggctagtccg tttttagcgc gtgcgccaat
      481 tctgcagaca aatggctcta gagagggcct atttcccatg attccttcat atttgcatat
      541 acgatacaag gctgttagag agataattgg aattaatttg actgtaaaca caaagatatt
      601 agtacaaaat acgtgacgta gaaagtaata atttcttggg tagtttgcag ttttaaaatt
      661 atgttttaaa atggactatc atatgcttac cgtaacttga aagtatttcg atttcttggc
      721 tttatatatc ttgtggaaag gacgaaacac cggagaccta ttcgccttaa ggtctcggtt
      781 ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtggc
      841 accgagtcgg tgcttttttg ttttagagct agaaatagca agttaaaata aggctagtcc
      901 gtttttagcg cgtgcgccaa ttctgcagac aaatggggta cccgttacat aacttacggt
      961 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa tagtaacgcc
     1021 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     1081 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
     1141 gcccgcctgg cattgtgccc agtacatgac cttatgggac tttcctactt ggcagtacat
     1201 ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct
     1261 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg
     1321 tgcagcgatg ggggcggggg gggggggggg gcgcgcgcca ggcggggcgg ggcggggcga
     1381 ggggcggggc ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg
     1441 aaagtttcct tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg
     1501 gcgggcggga gtcgctgcgc gctgccttcg ccccgtgccc cgctccgccg ccgcctcgcg
     1561 ccgcccgccc cggctctgac tgaccgcgtt actcccacag gtgagcgggc gggacggccc
     1621 ttctcctccg ggctgtaatt agctgagcaa gaggtaaggg tttaagggat ggttggttgg
     1681 tggggtatta atgtttaatt acctggagca cctgcctgaa atcacttttt ttcaggttgg
     1741 accggtgcca ccatggacta taaggaccac gacggagact acaaggatca tgatattgat
     1801 tacaaagacg atgacgataa gatggcccca aagaagaagc ggaaggtcgg tatccacgga
     1861 gtcccagcag ccgacaagaa gtacagcatc ggcctggaca tcggcaccaa ctctgtgggc
     1921 tgggccgtga tcaccgacga gtacaaggtg cccagcaaga aattcaaggt gctgggcaac
     1981 accgaccggc acagcatcaa gaagaacctg atcggagccc tgctgttcga cagcggcgaa
     2041 acagccgagg ccacccggct gaagagaacc gccagaagaa gatacaccag acggaagaac
     2101 cggatctgct atctgcaaga gatcttcagc aacgagatgg ccaaggtgga cgacagcttc
     2161 ttccacagac tggaagagtc cttcctggtg gaagaggata agaagcacga gcggcacccc
     2221 atcttcggca acatcgtgga cgaggtggcc taccacgaga agtaccccac catctaccac
     2281 ctgagaaaga aactggtgga cagcaccgac aaggccgacc tgcggctgat ctatctggcc
     2341 ctggcccaca tgatcaagtt ccggggccac ttcctgatcg agggcgacct gaaccccgac
     2401 aacagcgacg tggacaagct gttcatccag ctggtgcaga cctacaacca gctgttcgag
     2461 gaaaacccca tcaacgccag cggcgtggac gccaaggcca tcctgtctgc cagactgagc
     2521 aagagcagac ggctggaaaa tctgatcgcc cagctgcccg gcgagaagaa gaatggcctg
     2581 ttcggaaacc tgattgccct gagcctgggc ctgaccccca acttcaagag caacttcgac
     2641 ctggccgagg atgccaaact gcagctgagc aaggacacct acgacgacga cctggacaac
     2701 ctgctggccc agatcggcga ccagtacgcc gacctgtttc tggccgccaa gaacctgtcc
     2761 gacgccatcc tgctgagcga catcctgaga gtgaacaccg agatcaccaa ggcccccctg
     2821 agcgcctcta tgatcaagag atacgacgag caccaccagg acctgaccct gctgaaagct
     2881 ctcgtgcggc agcagctgcc tgagaagtac aaagagattt tcttcgacca gagcaagaac
     2941 ggctacgccg gctacattga cggcggagcc agccaggaag agttctacaa gttcatcaag
     3001 cccatcctgg aaaagatgga cggcaccgag gaactgctcg tgaagctgaa cagagaggac
     3061 ctgctgcgga agcagcggac cttcgacaac ggcagcatcc cccaccagat ccacctggga
     3121 gagctgcacg ccattctgcg gcggcaggaa gatttttacc cattcctgaa ggacaaccgg
     3181 gaaaagatcg agaagatcct gaccttccgc atcccctact acgtgggccc tctggccagg
     3241 ggaaacagca gattcgcctg gatgaccaga aagagcgagg aaaccatcac cccctggaac
     3301 ttcgaggaag tggtggacaa gggcgcttcc gcccagagct tcatcgagcg gatgaccaac
     3361 ttcgataaga acctgcccaa cgagaaggtg ctgcccaagc acagcctgct gtacgagtac
     3421 ttcaccgtgt ataacgagct gaccaaagtg aaatacgtga ccgagggaat gagaaagccc
     3481 gccttcctga gcggcgagca gaaaaaggcc atcgtggacc tgctgttcaa gaccaaccgg
     3541 aaagtgaccg tgaagcagct gaaagaggac tacttcaaga aaatcgagtg cttcgactcc
     3601 gtggaaatct ccggcgtgga agatcggttc aacgcctccc tgggcacata ccacgatctg
     3661 ctgaaaatta tcaaggacaa ggacttcctg gacaatgagg aaaacgagga cattctggaa
     3721 gatatcgtgc tgaccctgac actgtttgag gacagagaga tgatcgagga acggctgaaa
     3781 acctatgccc acctgttcga cgacaaagtg atgaagcagc tgaagcggcg gagatacacc
     3841 ggctggggca ggctgagccg gaagctgatc aacggcatcc gggacaagca gtccggcaag
     3901 acaatcctgg atttcctgaa gtccgacggc ttcgccaaca gaaacttcat gcagctgatc
     3961 cacgacgaca gcctgacctt taaagaggac atccagaaag cccaggtgtc cggccagggc
     4021 gatagcctgc acgagcacat tgccaatctg gccggcagcc ccgccattaa gaagggcatc
     4081 ctgcagacag tgaaggtggt ggacgagctc gtgaaagtga tgggccggca caagcccgag
     4141 aacatcgtga tcgaaatggc cagagagaac cagaccaccc agaagggaca gaagaacagc
     4201 cgcgagagaa tgaagcggat cgaagagggc atcaaagagc tgggcagcca gatcctgaaa
     4261 gaacaccccg tggaaaacac ccagctgcag aacgagaagc tgtacctgta ctacctgcag
     4321 aatgggcggg atatgtacgt ggaccaggaa ctggacatca accggctgtc cgactacgat
     4381 gtggaccata tcgtgcctca gagctttctg aaggacgact ccatcgacaa caaggtgctg
     4441 accagaagcg acaagaaccg gggcaagagc gacaacgtgc cctccgaaga ggtcgtgaag
     4501 aagatgaaga actactggcg gcagctgctg aacgccaagc tgattaccca gagaaagttc
     4561 gacaatctga ccaaggccga gagaggcggc ctgagcgaac tggataaggc cggcttcatc
     4621 aagagacagc tggtggaaac ccggcagatc acaaagcacg tggcacagat cctggactcc
     4681 cggatgaaca ctaagtacga cgagaatgac aagctgatcc gggaagtgaa agtgatcacc
     4741 ctgaagtcca agctggtgtc cgatttccgg aaggatttcc agttttacaa agtgcgcgag
     4801 atcaacaact accaccacgc ccacgacgcc tacctgaacg ccgtcgtggg aaccgccctg
     4861 atcaaaaagt accctaagct ggaaagcgag ttcgtgtacg gcgactacaa ggtgtacgac
     4921 gtgcggaaga tgatcgccaa gagcgagcag gaaatcggca aggctaccgc caagtacttc
     4981 ttctacagca acatcatgaa ctttttcaag accgagatta ccctggccaa cggcgagatc
     5041 cggaagcggc ctctgatcga gacaaacggc gaaaccgggg agatcgtgtg ggataagggc
     5101 cgggattttg ccaccgtgcg gaaagtgctg agcatgcccc aagtgaatat cgtgaaaaag
     5161 accgaggtgc agacaggcgg cttcagcaaa gagtctatcc tgcccaagag gaacagcgat
     5221 aagctgatcg ccagaaagaa ggactgggac cctaagaagt acggcggctt cgacagcccc
     5281 accgtggcct attctgtgct ggtggtggcc aaagtggaaa agggcaagtc caagaaactg
     5341 aagagtgtga aagagctgct ggggatcacc atcatggaaa gaagcagctt cgagaagaat
     5401 cccatcgact ttctggaagc caagggctac aaagaagtga aaaaggacct gatcatcaag
     5461 ctgcctaagt actccctgtt cgagctggaa aacggccgga agagaatgct ggcctctgcc
     5521 ggcgaactgc agaagggaaa cgaactggcc ctgccctcca aatatgtgaa cttcctgtac
     5581 ctggccagcc actatgagaa gctgaagggc tcccccgagg ataatgagca gaaacagctg
     5641 tttgtggaac agcacaagca ctacctggac gagatcatcg agcagatcag cgagttctcc
     5701 aagagagtga tcctggccga cgctaatctg gacaaagtgc tgtccgccta caacaagcac
     5761 cgggataagc ccatcagaga gcaggccgag aatatcatcc acctgtttac cctgaccaat
     5821 ctgggagccc ctgccgcctt caagtacttt gacaccacca tcgaccggaa gaggtacacc
     5881 agcaccaaag aggtgctgga cgccaccctg atccaccaga gcatcaccgg cctgtacgag
     5941 acacggatcg acctgtctca gctgggaggc gacaaaaggc cggcggccac gaaaaaggcc
     6001 ggccaggcaa aaaagaaaaa ggaattcgga tctcgcgagg gcagaggaag ccttctaaca
     6061 tgcggtgacg tggaggagaa tcccggccct atgaccgagt acaagcccac ggtgcgcctc
     6121 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac
     6181 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa
     6241 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc
     6301 gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag
     6361 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa
     6421 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc
     6481 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg
     6541 gccgagcgcg ccggggtgcc cgccttcctg gagacatccg cgccccgcaa cctccccttc
     6601 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc
     6661 tggtgcatga cccgcaagcc cggtgccgga tccggcgcaa caaacttctc tctgctgaaa
     6721 caagccggag atgtcgaaga gaatcctgga ccgatggtgt ctaagggcga agagctgttc
     6781 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
     6841 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
     6901 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
     6961 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
     7021 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
     7081 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     7141 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac
     7201 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc
     7261 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     7321 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc
     7381 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     7441 atcactctcg gcatggacga gctgtacaag tgagaattcc tagagctcgc tgatcagcct
     7501 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     7561 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     7621 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     7681 attgggaaga gaatagcagg catgctgggg agcggccgca ggaaccccta gtgatggagt
     7741 tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc
     7801 gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg
     7861 ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc
     7921 aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     7981 gcgcagcgtg accgctacac ttgccagcgc cttagcgccc gctcctttcg ctttcttccc
     8041 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     8101 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg
     8161 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     8221 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaactcta tctcgggcta
     8281 ttcttttgat ttataaggga ttttgccgat ttcggtctat tggttaaaaa atgagctgat
     8341 ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac
     8401 tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc
     8461 cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac
     8521 cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg
     8581 aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta
     8641 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     8701 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     8761 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     8821 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     8881 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     8941 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     9001 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     9061 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     9121 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     9181 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     9241 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     9301 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     9361 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     9421 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     9481 cggtgagcgt ggaagccgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     9541 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     9601 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     9661 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     9721 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     9781 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     9841 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     9901 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     9961 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
    10021 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
    10081 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
    10141 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
    10201 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
    10261 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
    10321 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
    10381 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgct
//
