LOCUS       pFH32                   6451 bp    DNA     circular UNA 10-APR-2019
DEFINITION  Level0 SpCas9-NG, human codon optimized.
ACCESSION   urn.local...12t-abnbxno
VERSION     urn.local...12t-abnbxno
KEYWORDS    .
SOURCE      
  ORGANISM  
            .
FEATURES             Location/Qualifiers
     CDS             215..1225
                     /label="Sm/Sp no DraIII"
     rep_origin      1385..1999
                     /modified_by="hahnf"
                     /label="rep (pMB1)"
     rep_origin      1984
                     /label="ORI"
     misc_feature    2225..2230
                     /Recognition_pattern="GGTCTC(1/5)"
                     /label="BsaI"
                     /note="Geneious type: restriction site"
     ligation        2232..2235
                     /label="Ligation"
     CDS             2233..6432
                     /created_by="hahnf"
                     /modified_by="hahnf"
                     /label="SpCas9-NG"
     sig_peptide     2233..2283
                     /created_by="hahnf"
                     /modified_by="hahnf"
                     /label="SV40 NLS"
     sig_peptide     6385..6432
                     /created_by="hahnf"
                     /modified_by="hahnf"
                     /label="Nucleoplasmin NLS"
     Stop_codon      6433..6435
                     /created_by="hahnf"
                     /label="Stop"
     ligation        6436..6439
                     /label="Ligation"
     misc_feature    complement(6441..6446)
                     /Recognition_pattern="GGTCTC(1/5)"
                     /label="BsaI"
                     /note="Geneious type: restriction site"
ORIGIN      
        1 gtggctcttc agtggacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
       61 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
      121 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
      181 ataaatgctt caataatatt gaaaaaggaa gagtatgcgc tcacgcaact ggtccagaac
      241 cttgaccgaa cgcagcggtg gtaacggcgc agtggcggtt ttcatggctt gttatgactg
      301 tttttttggg gtacagtcta tgcctcgggc atccaagcag caagcgcgtt acgccgtggg
      361 tcgatgtttg atgttatgga gcagcaacga tgttacgcag cagggcagtc gccctaaaac
      421 aaagttaaac atcatgaggg aagcggtgat cgccgaagta tcgactcaac tatcagaggt
      481 agttggcgtc atcgagcgcc atctcgaacc gacgttgctg gccgtacatt tgtacggctc
      541 cgcagtggat ggcggcctga agccacacag cgatattgat ttgctggtta cggtgaccgt
      601 aaggcttgat gaaacaacgc ggcgagcttt gatcaacgac cttttggaaa cttcggcttc
      661 ccctggagag agcgagattc tccgcgctgt agaagtcacc attgttgtgc acgacgacat
      721 cattccgtgg cgttatccag ctaagcgcga actgcaattt ggagaatggc agcgcaatga
      781 cattcttgca ggtatcttcg agccagccac gatcgacatt gatctggcta tcttgctgac
      841 aaaagcaaga gaacatagcg ttgccttggt aggtccagcg gcggaggaac tctttgatcc
      901 ggttcctgaa caggatctat ttgaggcgct aaatgaaacc ttaacgctat ggaactcgcc
      961 gcccgactgg gctggcgatg agcgaaatgt agtgcttacg ttgtcccgca tttggtacag
     1021 cgcagtaacc ggcaaaatcg cgccgaagga tgtcgctgcc gactgggcaa tggagcgcct
     1081 gccggcccag tatcagcccg tcatacttga agctagacag gcttatcttg gacaagaaga
     1141 agatcgcttg gcctcgcgcg cagatcagtt ggaagaattt gtccattacg taaaaggcga
     1201 gatcaccaag gtagtcggca aataactgtc agaccaagtt tactcatata tactttagat
     1261 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     1321 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     1381 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     1441 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     1501 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
     1561 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     1621 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     1681 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     1741 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     1801 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     1861 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     1921 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     1981 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     2041 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     2101 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     2161 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatcact
     2221 ctgtggtctc aaatggcccc aaagaagaag cggaaggtcg gtatccacgg agtcccagca
     2281 gccgacaaga agtacagcat cggcctggac atcggcacca actctgtggg ctgggccgtg
     2341 atcaccgacg agtacaaggt gcccagcaag aaattcaagg tgctgggcaa caccgaccgg
     2401 cacagcatca agaagaacct gatcggagcc ctgctgttcg acagcggcga aacagccgag
     2461 gccacccggc tgaagagaac cgccagaaga agatacacca gacggaagaa ccggatctgc
     2521 tatctgcaag agatcttcag caacgagatg gccaaggtgg acgacagctt cttccacaga
     2581 ctggaagagt ccttcctggt ggaagaggat aagaagcacg agcggcaccc catcttcggc
     2641 aacatcgtgg acgaggtggc ctaccacgag aagtacccca ccatctacca cctgagaaag
     2701 aaactggtgg acagcaccga caaggccgac ctgcggctga tctatctggc cctggcccac
     2761 atgatcaagt tccggggcca cttcctgatc gagggcgacc tgaaccccga caacagcgac
     2821 gtggacaagc tgttcatcca gctggtgcag acctacaacc agctgttcga ggaaaacccc
     2881 atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg ccagactgag caagagcaga
     2941 cggctggaaa atctgatcgc ccagctgccc ggcgagaaga agaatggcct gttcggaaac
     3001 ctgattgccc tgagcctggg cctgaccccc aacttcaaga gcaacttcga cctggccgag
     3061 gatgccaaac tgcagctgag caaggacacc tacgacgacg acctggacaa cctgctggcc
     3121 cagatcggcg accagtacgc cgacctgttt ctggccgcca agaacctgtc cgacgccatc
     3181 ctgctgagcg acatcctgag agtgaacacc gagatcacca aggcccccct gagcgcctct
     3241 atgatcaaga gatacgacga gcaccaccag gacctgaccc tgctgaaagc tctcgtgcgg
     3301 cagcagctgc ctgagaagta caaagagatt ttcttcgacc agagcaagaa cggctacgcc
     3361 ggctacattg acggcggagc cagccaggaa gagttctaca agttcatcaa gcccatcctg
     3421 gaaaagatgg acggcaccga ggaactgctc gtgaagctga acagagagga cctgctgcgg
     3481 aagcagcgga ccttcgacaa cggcagcatc ccccaccaga tccacctggg agagctgcac
     3541 gccattctgc ggcggcagga agatttttac ccattcctga aggacaaccg ggaaaagatc
     3601 gagaagatcc tgaccttccg catcccctac tacgtgggcc ctctggccag gggaaacagc
     3661 agattcgcct ggatgaccag aaagagcgag gaaaccatca ccccctggaa cttcgaggaa
     3721 gtggtggaca agggcgcttc cgcccagagc ttcatcgagc ggatgaccaa cttcgataag
     3781 aacctgccca acgagaaggt gctgcccaag cacagcctgc tgtacgagta cttcaccgtg
     3841 tataacgagc tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc cgccttcctg
     3901 agcggcgagc agaaaaaggc catcgtggac ctgctgttca agaccaaccg gaaagtgacc
     3961 gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc
     4021 tccggcgtgg aagatcggtt caacgcctcc ctgggcacat accacgatct gctgaaaatt
     4081 atcaaggaca aggacttcct ggacaatgag gaaaacgagg acattctgga agatatcgtg
     4141 ctgaccctga cactgtttga ggacagagag atgatcgagg aacggctgaa aacctatgcc
     4201 cacctgttcg acgacaaagt gatgaagcag ctgaagcggc ggagatacac cggctggggc
     4261 aggctgagcc ggaagctgat caacggcatc cgggacaagc agtccggcaa gacaatcctg
     4321 gatttcctga agtccgacgg cttcgccaac agaaacttca tgcagctgat ccacgacgac
     4381 agcctgacct ttaaagagga catccagaaa gcccaggtgt ccggccaggg cgatagcctg
     4441 cacgagcaca ttgccaatct ggccggcagc cccgccatta agaagggcat cctgcagaca
     4501 gtgaaggtgg tggacgagct cgtgaaagtg atgggccggc acaagcccga gaacatcgtg
     4561 atcgaaatgg ccagagagaa ccagaccacc cagaagggac agaagaacag ccgcgagaga
     4621 atgaagcgga tcgaagaggg catcaaagag ctgggcagcc agatcctgaa agaacacccc
     4681 gtggaaaaca cccagctgca gaacgagaag ctgtacctgt actacctgca gaatgggcgg
     4741 gatatgtacg tggaccagga actggacatc aaccggctgt ccgactacga tgtggaccat
     4801 atcgtgcctc agagctttct gaaggacgac tccatcgaca acaaggtgct gaccagaagc
     4861 gacaagaacc ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag
     4921 aactactggc ggcagctgct gaacgccaag ctgattaccc agagaaagtt cgacaatctg
     4981 accaaggccg agagaggcgg cctgagcgaa ctggataagg ccggcttcat caagagacag
     5041 ctggtggaaa cccggcagat cacaaagcac gtggcacaga tcctggactc ccggatgaac
     5101 actaagtacg acgagaatga caagctgatc cgggaagtga aagtgatcac cctgaagtcc
     5161 aagctggtgt ccgatttccg gaaggatttc cagttttaca aagtgcgcga gatcaacaac
     5221 taccaccacg cccacgacgc ctacctgaac gccgtcgtgg gaaccgccct gatcaaaaag
     5281 taccctaagc tggaaagcga gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag
     5341 atgatcgcca agagcgagca ggaaatcggc aaggctaccg ccaagtactt cttctacagc
     5401 aacatcatga actttttcaa gaccgagatt accctggcca acggcgagat ccggaagcgg
     5461 cctctgatcg agacaaacgg cgaaaccggg gagatcgtgt gggataaggg ccgggatttt
     5521 gccaccgtgc ggaaagtgct gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg
     5581 cagacaggcg gcttcagcaa agagtctatc cggcccaaga ggaacagcga taagctgatc
     5641 gccagaaaga aggactggga ccctaagaag tacggcggct tcgttagccc caccgtggcc
     5701 tattctgtgc tggtggtggc caaagtggaa aagggcaagt ccaagaaact gaagagtgtg
     5761 aaagagctgc tggggatcac catcatggaa agaagcagct tcgagaagaa tcccatcgac
     5821 tttctggaag ccaagggcta caaagaagtg aaaaaggacc tgatcatcaa gctgcctaag
     5881 tactccctgt tcgagctgga aaacggccgg aagagaatgc tggcctctgc ccggttcctg
     5941 cagaagggaa acgaactggc cctgccctcc aaatatgtga acttcctgta cctggccagc
     6001 cactatgaga agctgaaggg ctcccccgag gataatgagc agaaacagct gtttgtggaa
     6061 cagcacaagc actacctgga cgagatcatc gagcagatca gcgagttctc caagagagtg
     6121 atcctggccg acgctaatct ggacaaagtg ctgtccgcct acaacaagca ccgggataag
     6181 cccatcagag agcaggccga gaatatcatc cacctgttta ccctgaccaa tctgggagcc
     6241 cctcgggcct tcaagtactt tgacaccacc atcgaccgga aggtgtaccg gagcaccaaa
     6301 gaggtgctgg acgccaccct gatccaccag agcatcaccg gcctgtacga gacacggatc
     6361 gacctgtctc agctgggagg cgacaaaagg ccggcggcca cgaaaaaggc cggccaggca
     6421 aaaaagaaaa agtaagcttt gagaccacga a
//
