LOCUS       .                       8947 bp    DNA     circular UNK 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   
VERSION     
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     intron          208..435
                     /label="hybrid intron"
                     /note="hybrid between chicken β-actin (CBA) and minute
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             573..4673
                     /label="Cas9"
                     /note="S. pyogenes Cas9, codon optimized for C. elegans
                     with synthetic introns"
     polyA_signal    4755..4962
                     /label="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4971..5111
                     /label="AAV2 ITR"
                     /note="inverted terminal repeat of adeno-associated virus
                     serotype 2"
     rep_origin      5186..5641
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        5923..6027
                     /label="AmpR promoter"
     CDS             6028..6885
                     /label="AmpR"
                     /note="β-lactamase"
     rep_origin      7059..7647
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     misc_RNA        7976..8051
                     /label="gRNA scaffold"
                     /note="guide RNA scaffold for the Streptococcus pyogenes
                     CRISPR/Cas9 system"
     promoter        8149..8389
                     /label="U6 promoter"
                     /note="RNA polymerase III promoter for human U6 snRNA"
     enhancer        8590..8875
                     /label="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer;
                     contains an 18-bp deletion relative to the standard CMV
                     enhancer"
     promoter        8877..9154
                     /label="chicken β-actin promoter"
ORIGIN
        1 tttatttttt aattattttg tgcagcgatg ggggcggggg gggggggggg gcgcgcgcca
       61 ggcggggcgg ggcggggcga ggggcggggc ggggcgaggc ggagaggtgc ggcggcagcc
      121 aatcagagcg gcgcgctccg aaagtttcct tttatggcga ggcggcggcg gcggcggccc
      181 tataaaaagc gaagcgcgcg gcgggcggga gtcgctgcgc gctgccttcg ccccgtgccc
      241 cgctccgccg ccgcctcgcg ccgcccgccc cggctctgac tgaccgcgtt actcccacag
      301 gtgagcgggc gggacggccc ttctcctccg ggctgtaatt agctgagcaa gaggtaaggg
      361 tttaagggat ggttggttgg tggggtatta atgtttaatt acctggagca cctgcctgaa
      421 atcacttttt ttcaggttgg accggtgcca ccatggacta taaggaccac gacggagact
      481 acaaggatca tgatattgat tacaaagacg atgacgataa gatggcccca aagaagaagc
      541 ggaaggtcgg tatccacgga gtcccagcag ccgacaagaa gtacagcatc ggcctggaca
      601 tcggcaccaa ctctgtgggc tgggccgtga tcaccgacga gtacaaggtg cccagcaaga
      661 aattcaaggt gctgggcaac accgaccggc acagcatcaa gaagaacctg atcggagccc
      721 tgctgttcga cagcggcgaa acagccgagg ccacccggct gaagagaacc gccagaagaa
      781 gatacaccag acggaagaac cggatctgct atctgcaaga gatcttcagc aacgagatgg
      841 ccaaggtgga cgacagcttc ttccacagac tggaagagtc cttcctggtg gaagaggata
      901 agaagcacga gcggcacccc atcttcggca acatcgtgga cgaggtggcc taccacgaga
      961 agtaccccac catctaccac ctgagaaaga aactggtgga cagcaccgac aaggccgacc
     1021 tgcggctgat ctatctggcc ctggcccaca tgatcaagtt ccggggccac ttcctgatcg
     1081 agggcgacct gaaccccgac aacagcgacg tggacaagct gttcatccag ctggtgcaga
     1141 cctacaacca gctgttcgag gaaaacccca tcaacgccag cggcgtggac gccaaggcca
     1201 tcctgtctgc cagactgagc aagagcagac ggctggaaaa tctgatcgcc cagctgcccg
     1261 gcgagaagaa gaatggcctg ttcggaaacc tgattgccct gagcctgggc ctgaccccca
     1321 acttcaagag caacttcgac ctggccgagg atgccaaact gcagctgagc aaggacacct
     1381 acgacgacga cctggacaac ctgctggccc agatcggcga ccagtacgcc gacctgtttc
     1441 tggccgccaa gaacctgtcc gacgccatcc tgctgagcga catcctgaga gtgaacaccg
     1501 agatcaccaa ggcccccctg agcgcctcta tgatcaagag atacgacgag caccaccagg
     1561 acctgaccct gctgaaagct ctcgtgcggc agcagctgcc tgagaagtac aaagagattt
     1621 tcttcgacca gagcaagaac ggctacgccg gctacattga cggcggagcc agccaggaag
     1681 agttctacaa gttcatcaag cccatcctgg aaaagatgga cggcaccgag gaactgctcg
     1741 tgaagctgaa cagagaggac ctgctgcgga agcagcggac cttcgacaac ggcagcatcc
     1801 cccaccagat ccacctggga gagctgcacg ccattctgcg gcggcaggaa gatttttacc
     1861 cattcctgaa ggacaaccgg gaaaagatcg agaagatcct gaccttccgc atcccctact
     1921 acgtgggccc tctggccagg ggaaacagca gattcgcctg gatgaccaga aagagcgagg
     1981 aaaccatcac cccctggaac ttcgaggaag tggtggacaa gggcgcttcc gcccagagct
     2041 tcatcgagcg gatgaccaac ttcgataaga acctgcccaa cgagaaggtg ctgcccaagc
     2101 acagcctgct gtacgagtac ttcaccgtgt ataacgagct gaccaaagtg aaatacgtga
     2161 ccgagggaat gagaaagccc gccttcctga gcggcgagca gaaaaaggcc atcgtggacc
     2221 tgctgttcaa gaccaaccgg aaagtgaccg tgaagcagct gaaagaggac tacttcaaga
     2281 aaatcgagtg cttcgactcc gtggaaatct ccggcgtgga agatcggttc aacgcctccc
     2341 tgggcacata ccacgatctg ctgaaaatta tcaaggacaa ggacttcctg gacaatgagg
     2401 aaaacgagga cattctggaa gatatcgtgc tgaccctgac actgtttgag gacagagaga
     2461 tgatcgagga acggctgaaa acctatgccc acctgttcga cgacaaagtg atgaagcagc
     2521 tgaagcggcg gagatacacc ggctggggca ggctgagccg gaagctgatc aacggcatcc
     2581 gggacaagca gtccggcaag acaatcctgg atttcctgaa gtccgacggc ttcgccaaca
     2641 gaaacttcat gcagctgatc cacgacgaca gcctgacctt taaagaggac atccagaaag
     2701 cccaggtgtc cggccagggc gatagcctgc acgagcacat tgccaatctg gccggcagcc
     2761 ccgccattaa gaagggcatc ctgcagacag tgaaggtggt ggacgagctc gtgaaagtga
     2821 tgggccggca caagcccgag aacatcgtga tcgaaatggc cagagagaac cagaccaccc
     2881 agaagggaca gaagaacagc cgcgagagaa tgaagcggat cgaagagggc atcaaagagc
     2941 tgggcagcca gatcctgaaa gaacaccccg tggaaaacac ccagctgcag aacgagaagc
     3001 tgtacctgta ctacctgcag aatgggcggg atatgtacgt ggaccaggaa ctggacatca
     3061 accggctgtc cgactacgat gtggaccata tcgtgcctca gagctttctg aaggacgact
     3121 ccatcgacaa caaggtgctg accagaagcg acaagaaccg gggcaagagc gacaacgtgc
     3181 cctccgaaga ggtcgtgaag aagatgaaga actactggcg gcagctgctg aacgccaagc
     3241 tgattaccca gagaaagttc gacaatctga ccaaggccga gagaggcggc ctgagcgaac
     3301 tggataaggc cggcttcatc aagagacagc tggtggaaac ccggcagatc acaaagcacg
     3361 tggcacagat cctggactcc cggatgaaca ctaagtacga cgagaatgac aagctgatcc
     3421 gggaagtgaa agtgatcacc ctgaagtcca agctggtgtc cgatttccgg aaggatttcc
     3481 agttttacaa agtgcgcgag atcaacaact accaccacgc ccacgacgcc tacctgaacg
     3541 ccgtcgtggg aaccgccctg atcaaaaagt accctaagct ggaaagcgag ttcgtgtacg
     3601 gcgactacaa ggtgtacgac gtgcggaaga tgatcgccaa gagcgagcag gaaatcggca
     3661 aggctaccgc caagtacttc ttctacagca acatcatgaa ctttttcaag accgagatta
     3721 ccctggccaa cggcgagatc cggaagcggc ctctgatcga gacaaacggc gaaaccgggg
     3781 agatcgtgtg ggataagggc cgggattttg ccaccgtgcg gaaagtgctg agcatgcccc
     3841 aagtgaatat cgtgaaaaag accgaggtgc agacaggcgg cttcagcaaa gagtctatcc
     3901 tgcccaagag gaacagcgat aagctgatcg ccagaaagaa ggactgggac cctaagaagt
     3961 acggcggctt cgacagcccc accgtggcct attctgtgct ggtggtggcc aaagtggaaa
     4021 agggcaagtc caagaaactg aagagtgtga aagagctgct ggggatcacc atcatggaaa
     4081 gaagcagctt cgagaagaat cccatcgact ttctggaagc caagggctac aaagaagtga
     4141 aaaaggacct gatcatcaag ctgcctaagt actccctgtt cgagctggaa aacggccgga
     4201 agagaatgct ggcctctgcc ggcgaactgc agaagggaaa cgaactggcc ctgccctcca
     4261 aatatgtgaa cttcctgtac ctggccagcc actatgagaa gctgaagggc tcccccgagg
     4321 ataatgagca gaaacagctg tttgtggaac agcacaagca ctacctggac gagatcatcg
     4381 agcagatcag cgagttctcc aagagagtga tcctggccga cgctaatctg gacaaagtgc
     4441 tgtccgccta caacaagcac cgggataagc ccatcagaga gcaggccgag aatatcatcc
     4501 acctgtttac cctgaccaat ctgggagccc ctgccgcctt caagtacttt gacaccacca
     4561 tcgaccggaa gaggtacacc agcaccaaag aggtgctgga cgccaccctg atccaccaga
     4621 gcatcaccgg cctgtacgag acacggatcg acctgtctca gctgggaggc gacaaaaggc
     4681 cggcggccac gaaaaaggcc ggccaggcaa aaaagaaaaa gtaagaattc ctagagctcg
     4741 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     4801 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     4861 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     4921 caagggggag gattgggaag agaatagcag gcatgctggg gagcggccgc aggaacccct
     4981 agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc
     5041 aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag
     5101 ctgcctgcag gggcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca
     5161 ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt
     5221 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccttagcgcc cgctcctttc
     5281 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     5341 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     5401 ttgggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     5461 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaactct
     5521 atctcgggct attcttttga tttataaggg attttgccga tttcggtcta ttggttaaaa
     5581 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt
     5641 ttatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
     5701 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     5761 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     5821 cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
     5881 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     5941 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     6001 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     6061 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     6121 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     6181 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     6241 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     6301 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     6361 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     6421 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     6481 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     6541 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     6601 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     6661 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     6721 aaatctggag ccggtgagcg tggaagccgc ggtatcattg cagcactggg gccagatggt
     6781 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     6841 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     6901 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     6961 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     7021 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     7081 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     7141 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     7201 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     7261 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     7321 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     7381 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     7441 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     7501 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     7561 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     7621 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     7681 gccttttgct ggccttttgc tcacatgtga gggcctattt cccatgattc cttcatattt
     7741 gcatatacga tacaaggctg ttagagagat aattggaatt aatttgactg taaacacaaa
     7801 gatattagta caaaatacgt gacgtagaaa gtaataattt cttgggtagt ttgcagtttt
     7861 aaaattatgt tttaaaatgg actatcatat gcttaccgta acttgaaagt atttcgattt
     7921 cttggcttta tatatcttgt ggaaaggacg aaacaccggg tcttcgagaa gacctgtttt
     7981 agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac
     8041 cgagtcggtg cttttttgtt ttagagctag aaatagcaag ttaaaataag gctagtccgt
     8101 ttttagcgcg tgcgccaatt ctgcagacaa atggctctag aggcatgtga gggcctattt
     8161 cccatgattc cttcatattt gcatatacga tacaaggctg ttagagagat aattggaatt
     8221 aatttgactg taaacacaaa gatattagta caaaatacgt gacgtagaaa gtaataattt
     8281 cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg actatcatat gcttaccgta
     8341 acttgaaagt atttcgattt cttggcttta tatatcttgt ggaaaggacg aaacaccgca
     8401 tcgtacgcgt acgtgttgtt ttagagctag aaatagcaag ttaaaataag gctagtccgt
     8461 tatcaacttg aaaaagtggc accgagtcgg tgcttttttg ttttagagct agaaatagca
     8521 agttaaaata aggctagtcc gtttttagcg cgtgcgccaa ttctgcagac aaatggctct
     8581 agaggtaccc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     8641 ccgcccattg acgtcaatag taacgccaat agggactttc cattgacgtc aatgggtgga
     8701 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc
     8761 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tgtgcccagt acatgacctt
     8821 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtcga
     8881 ggtgagcccc acgttctgct tcactctccc catctccccc ccctccccac ccccaatttt
     8941 gtattta
//
