LOCUS       sequence-202507-       11837 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SpCas9 fused to dNmCas9 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11837)
  AUTHORS   Bolukbasi MF, Liu P, Luk K, Kwok SF, Gupta A, Amrani N, Sontheimer 
            EJ, Zhu LJ, Wolfe SA
  TITLE     Orthogonal Cas9-Cas9 chimeras provide a versatile platform for 
            genome editing.
  JOURNAL   Nat Commun. 2018 Nov 19;9(1):4856. doi: 10.1038/s41467-018-07310-x.
  PUBMED    30451839
REFERENCE   2  (bases 1 to 11837)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2018-11-19"; volume: "9(1):4856. doi"; pages: " 
            10.1038/s41467-018-07310-x"
COMMENT     Sequence Label: pCSD_2XNLS-SpCas9WT-NLS-3XHA-2XNLS-NmdCas9-NLS
FEATURES             Location/Qualifiers
     source          1..11837
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        37..55
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     37..54
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             105..152
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             171..191
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             198..4298
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             198..4298
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             198..4298
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             4311..4331
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4332..4421
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     CDS             4434..4460
                     /codon_start=1
                     /product="nuclear localization signal (Makkerh et al., 
                     1996)"
                     /label=NLS
                     /translation="PAAKKKKLD"
     CDS             4476..4496
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(4497..4538,4539..4541,4542..6251,6252..6257,6258..6323,
                     6324..6326,6327..7739)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Neisseria meningitidis Type II 
                     CRISPR/Cas system"
                     /label=NmdCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the mutations D16A, D587A, 
                     H588A, and N611A"
                     /translation="AAFKPNPINYILGLAIGIASVGWAMVEIDEDENPICLIDLGVRVF
                     ERAEVPKTGDSLAMARRLARSVRRLTRRRAHRLLRARRLLKREGVLQAADFDENGLIKS
                     LPNTPWQLRAAALDRKLTPLEWSAVLLHLIKHRGYLSQRKNEGETADKELGALLKGVAD
                     NAHALQTGDFRTPAELALNKFEKESGHIRNQRGDYSHTFSRKDLQAELILLFEKQKEFG
                     NPHVSGGLKEGIETLLMTQRPALSGDAVQKMLGHCTFEPAEPKAAKNTYTAERFIWLTK
                     LNNLRILEQGSERPLTDTERATLMDEPYRKSKLTYAQARKLLGLEDTAFFKGLRYGKDN
                     AEASTLMEMKAYHAISRALEKEGLKDKKSPLNLSPELQDEIGTAFSLFKTDEDITGRLK
                     DRIQPEILEALLKHISFDKFVQISLKALRRIVPLMEQGKRYDEACAEIYGDHYGKKNTE
                     EKIYLPPIPADEIRNPVVLRALSQARKVINGVVRRYGSPARIHIETAREVGKSFKDRKE
                     IEKRQEENRKDREKAAAKFREYFPNFVGEPKSKDILKLRLYEQQHGKCLYSGKEINLGR
                     LNEKGYVEIAAALPFSRTWDDSFNNKVLVLGSEAQNKGNQTPYEYFNGKDNSREWQEFK
                     ARVETSRFPRSKKQRILLQKFDEDGFKERNLNDTRYVNRFLCQFVADRMRLTGKGKKRV
                     FASNGQITNLLRGFWGLRKVRAENDRHHALDAVVVACSTVAMQQKITRFVRYKEMNAFD
                     GKTIDKETGEVLHQKTHFPQPWEFFAQEVMIRVFGKPDGKPEFEEADTPEKLRTLLAEK
                     LSSRPEAVHEYVTPLFVSRAPNRKMSGQGHMETVKSAKRLDEGVSVLRVPLTQLKLKDL
                     EKMVNREREPKLYEALKARLEAHKDDPAKAFAEPFYKYDKAGNRTQQVKAVRVEQVQKT
                     GVWVRNHNGIADNATMVRVDVFEKGDKYYLVPIYSWQVAKGILPDRAVVQGKDEEDWQL
                     IDDSFNFKFSLHPNDLVEVITKKARMFGYFASCHRGTGNINIRIHDLDHKIGKNGILEG
                     IGVKTALSFQKYQIDELGKEIRPCRLKKRPPVR"
     regulatory      7326..7335
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             7752..7772
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7788..7835
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     polyA_signal    7888..8022
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7912..7931)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     7966..7985
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(8135..8155)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(8135..8153)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(8174..8190)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8174..8190)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(8187..8209)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    8198..8214
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(8222..8228,8229..8246,8247..8252))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8267..8288
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8405..8422)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8576..9164)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8656..8675)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9335..10126,10127..10195))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     9958..9977
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(10196..10300)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(10326..10781)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10463..10484)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     10675..10694
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(10855..11837,1..2)
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 accgccattc tgcctgggga cgtcggagca agcttgattt aggtgacact atagaataca
       61 agctacttgt tctttttgca ggatcccatc gattcgaatt catgaaaagg ccggcggcca
      121 cgaaaaaggc cggccaggca aaaaagaaaa agctggacgg cagcggaagc cccaagaaga
      181 agaggaaggt accatgggac aagaagtact ccattgggct cgatatcggt accaacagcg
      241 tcggctgggc cgtcattacg gacgagtaca aggtgccgag caaaaaattc aaagttctgg
      301 gcaataccga tcgccacagc ataaagaaga acctcattgg agccctcctg ttcgactccg
      361 gggagacggc cgaagccacg cggctcaaaa gaacagcacg gcgcagatat acccgcagaa
      421 agaatcggat ctgctacctg caggagatct ttagtaatga gatggctaag gtggatgact
      481 ctttcttcca taggctggag gagtcctttt tggtggagga ggataaaaag cacgagcgcc
      541 acccaatctt tggcaatatc gtggacgagg tggcgtacca tgaaaagtac ccaaccatat
      601 atcatctgag gaagaagctg gtagacagta ctgataaggc tgacttgcgg ttgatctatc
      661 tcgcgctggc gcacatgatc aaatttcggg gacacttcct catcgagggg gacctgaacc
      721 cagacaacag cgatgtcgac aaactcttta tccaactggt tcagacttac aatcagcttt
      781 tcgaggagaa cccgatcaac gcatccggcg ttgacgccaa agcaatcctg agcgctaggc
      841 tgtccaaatc ccggcggctc gaaaacctca tcgcacagct ccctggggag aagaagaacg
      901 gcctgtttgg taatcttatc gccctgtcac tcgggctgac ccccaacttt aaatctaact
      961 tcgacctggc cgaagatgcc aagctgcaac tgagcaaaga cacctacgat gatgatctcg
     1021 acaatctgct ggcccagatc ggcgaccagt acgcagacct ttttttggcg gcaaagaacc
     1081 tgtcagacgc cattctgctg agtgatattc tgcgagtgaa cacggagatc accaaagctc
     1141 cgctgagcgc tagtatgatc aagcgctatg atgagcacca ccaagacttg actttgctga
     1201 aggcccttgt cagacagcaa ctgcctgaga agtacaagga aattttcttc gatcagtcta
     1261 aaaatggcta cgccggatac attgacggcg gagcaagcca ggaggaattt tacaaattta
     1321 ttaagcccat cttggaaaaa atggacggca ccgaggagct gctggtaaag ctgaacagag
     1381 aagatctgtt gcgcaaacag cgcactttcg acaatggaag catcccccac cagattcacc
     1441 tgggcgaact gcacgctatc ctcaggcggc aagaggattt ctaccccttt ttgaaagata
     1501 acagggaaaa gattgagaaa atcctcacat ttcggatacc ctactatgta ggccccctcg
     1561 ctcggggaaa ttccagattc gcgtggatga ctcgcaaatc agaagagacc atcactccct
     1621 ggaacttcga ggaagtcgtg gataaggggg cctctgccca gtccttcatc gaaaggatga
     1681 ctaactttga taaaaatctg cctaacgaaa aggtgcttcc taaacactct ctgctgtacg
     1741 agtacttcac agtttataac gagctcacca aggtcaaata cgtcacagaa gggatgagaa
     1801 agccagcatt cctgtctgga gagcagaaga aagctatcgt ggacctcctc ttcaagacga
     1861 accggaaagt taccgtgaaa cagctcaaag aagactattt caaaaagatt gaatgtttcg
     1921 actctgttga aatcagcgga gtggaggatc gcttcaacgc atccctggga acgtatcacg
     1981 atctcctgaa aatcattaaa gacaaggact tcctggacaa tgaggagaac gaggacattc
     2041 ttgaggacat tgtcctcacc cttacgttgt ttgaagatag ggagatgatt gaagaacgct
     2101 tgaaaactta cgctcatctc ttcgacgaca aagtcatgaa acagctcaag agacgccgat
     2161 atacaggatg ggggcggctg tcaagaaaac tgatcaatgg catccgagac aagcagagtg
     2221 gaaagacaat cctggatttt cttaagtccg atggatttgc caaccggaac ttcatgcagt
     2281 tgatccatga tgactctctc acctttaagg aggacatcca gaaagcacaa gtttctggcc
     2341 agggggacag tcttcacgag cacatcgcta atcttgcagg tagcccagct atcaaaaagg
     2401 gaatactgca gaccgttaag gtcgtggatg aactcgtcaa agtaatggga aggcataagc
     2461 ccgagaatat cgttatcgag atggcccgag agaaccaaac tacccagaag ggacagaaga
     2521 acagtaggga aaggatgaag aggattgaag agggtataaa agaactgggg tcccaaatcc
     2581 ttaaggaaca cccagttgaa aacacccagc ttcagaatga gaagctctac ctgtactacc
     2641 tgcagaacgg cagggacatg tacgtggatc aggaactgga catcaaccgg ttgtccgact
     2701 acgacgtgga tcatatcgtg ccccaaagct ttctcaaaga tgattctatt gataataaag
     2761 tgttgacaag atccgataaa aatagaggga agagtgataa cgtcccctca gaagaagttg
     2821 tcaagaaaat gaaaaattat tggcggcagc tgctgaacgc caaactgatc acacaacgga
     2881 agttcgataa tctgactaag gctgaacgag gtggcctgtc tgagttggat aaagccggct
     2941 tcatcaaaag gcagcttgtt gagacacgcc agatcaccaa gcacgtggcc caaattctcg
     3001 attcacgcat gaacaccaag tacgatgaaa atgacaaact gattcgagag gtgaaagtta
     3061 ttactctgaa gtctaagctg gtctcagatt tcagaaagga ctttcagttt tataaggtga
     3121 gagagatcaa caattaccac catgcgcatg atgcctacct gaatgcagtg gtaggcactg
     3181 cacttatcaa aaaatatccc aagctggaat ctgaatttgt ttacggagac tataaagtgt
     3241 acgatgttag gaaaatgatc gcaaagtctg agcaggaaat aggcaaggcc accgctaagt
     3301 acttctttta cagcaatatt atgaattttt tcaagaccga gattacactg gccaatggag
     3361 agattcggaa gcgaccactt atcgaaacaa acggagaaac aggagaaatc gtgtgggaca
     3421 agggtaggga tttcgcgaca gtccgcaagg tcctgtccat gccgcaggtg aacatcgtta
     3481 aaaagaccga agtacagacc ggaggcttct ccaaggaaag tatcctcccg aaaaggaaca
     3541 gcgacaagct gatcgcacgc aaaaaagatt gggaccccaa gaaatacggc ggattcgatt
     3601 ctcctacagt cgcttacagt gtactggttg tggccaaagt ggagaaaggg aagtctaaaa
     3661 aactcaaaag cgtcaaggaa ctgctgggca tcacaatcat ggagcgatcc agcttcgaga
     3721 aaaaccccat cgactttctc gaagcgaaag gatataaaga ggtcaaaaaa gacctcatca
     3781 ttaagctgcc caagtactct ctctttgagc ttgaaaacgg ccggaaacga atgctcgcta
     3841 gtgcgggcga gctgcagaaa ggtaacgagc tggcactgcc ctctaaatac gttaatttct
     3901 tgtatctggc cagccactat gaaaagctca aagggtctcc cgaagataat gagcagaagc
     3961 agctgttcgt ggaacaacac aaacactacc ttgatgagat catcgagcaa ataagcgagt
     4021 tctccaaaag agtgatcctc gccgacgcta acctcgataa ggtgctttct gcttacaata
     4081 agcacaggga taagcccatc agggagcagg cagaaaacat tatccacttg tttactctga
     4141 ccaacttggg cgcgcctgca gccttcaagt acttcgacac caccatagac agaaagcggt
     4201 acacctctac aaaggaggtc ctggacgcca cactgattca tcagtcaatt acggggctct
     4261 atgaaacaag aatcgacctc tctcagctcg gtggagacgg caccggcggg cccaagaaga
     4321 agaggaaggt atacccatac gatgttcctg actatgcggg ctatccctat gacgtcccgg
     4381 actatgcagg atcgtatcct tatgacgttc cagattacgc tggatccgcc gctccggcag
     4441 ctaagaaaaa gaaactggat ttcgaatccg gaatgcccaa gaagaagagg aaggtggccg
     4501 ccttcaagcc caaccccatc aactacatcc tgggcctggc catcggcatc gccagcgtgg
     4561 gctgggccat ggtggagatc gacgaggacg agaaccccat ctgcctgatc gacctgggtg
     4621 tgcgcgtgtt cgagcgcgct gaggtgccca agactggtga cagtctggct atggctcgcc
     4681 ggcttgctcg ctctgttcgg cgccttactc gccggcgcgc tcaccgcctt ctgcgcgctc
     4741 gccgcctgct gaagcgcgag ggtgtgctgc aggctgccga cttcgacgag aacggcctga
     4801 tcaagagcct gcccaacact ccttggcagc tgcgcgctgc cgctctggac cgcaagctga
     4861 ctcctctgga gtggagcgcc gtgctgctgc acctgatcaa gcaccgcggc tacctgagcc
     4921 agcgcaagaa cgagggcgag accgccgaca aggagctggg tgctctgctg aagggcgtgg
     4981 ccgacaacgc ccacgccctg cagactggtg acttccgcac tcctgctgag ctggccctga
     5041 acaagttcga gaaggagagc ggccacatcc gcaaccagcg cggcgactac agccacacct
     5101 tcagccgcaa ggacctgcag gccgagctga tcctgctgtt cgagaagcag aaggagttcg
     5161 gcaaccccca cgtgagcggc ggcctgaagg agggcatcga gaccctgctg atgacccagc
     5221 gccccgccct gagcggcgac gccgtgcaga agatgctggg ccactgcacc ttcgagccag
     5281 ccgagcccaa ggccgccaag aacacctaca ccgccgagcg cttcatctgg ctgaccaagc
     5341 tgaacaacct gcgcatcctg gagcagggca gcgagcgccc cctgaccgac accgagcgcg
     5401 ccaccctgat ggacgagccc taccgcaaga gcaagctgac ctacgcccag gcccgcaagc
     5461 tgctgggtct ggaggacacc gccttcttca agggcctgcg ctacggcaag gacaacgccg
     5521 aggccagcac cctgatggag atgaaggcct accacgccat cagccgcgcc ctggagaagg
     5581 agggcctgaa ggacaagaag agtcctctga acctgagccc cgagctgcag gacgagatcg
     5641 gcaccgcctt cagcctgttc aagaccgacg aggacatcac cggccgcctg aaggaccgca
     5701 tccagcccga gatcctggag gccctgctga agcacatcag cttcgacaag ttcgtgcaga
     5761 tcagcctgaa ggccctgcgc cgcatcgtgc ccctgatgga gcagggcaag cgctacgacg
     5821 aggcctgcgc cgagatctac ggcgaccact acggcaagaa gaacaccgag gagaagatct
     5881 acctgcctcc tatccccgcc gacgagatcc gcaaccccgt ggtgctgcgc gccctgagcc
     5941 aggcccgcaa ggtgatcaac ggcgtggtgc gccgctacgg cagccccgcc cgcatccaca
     6001 tcgagaccgc ccgcgaggtg ggcaagagct tcaaggaccg caaggagatc gagaagcgcc
     6061 aggaggagaa ccgcaaggac cgcgagaagg ccgccgccaa gttccgcgag tacttcccca
     6121 acttcgtggg cgagcccaag agcaaggaca tcctgaagct gcgcctgtac gagcagcagc
     6181 acggcaagtg cctgtacagc ggcaaggaga tcaacctggg ccgcctgaac gagaagggct
     6241 acgtggagat cgccgctgcc ctgcccttca gccgcacctg ggacgacagc ttcaacaaca
     6301 aggtgctggt gctgggcagc gaggctcaga acaagggcaa ccagaccccc tacgagtact
     6361 tcaacggcaa ggacaacagc cgcgagtggc aggagttcaa ggcccgcgtg gagaccagcc
     6421 gcttcccccg cagcaagaag cagcgcatcc tgctgcagaa gttcgacgag gacggcttca
     6481 aggagcgcaa cctgaacgac acccgctacg tgaaccgctt cctgtgccag ttcgtggccg
     6541 accgcatgcg cctgaccggc aagggcaaga agcgcgtgtt cgccagcaac ggccagatca
     6601 ccaacctgct gcgcggcttc tggggcctgc gcaaggtgcg cgccgagaac gaccgccacc
     6661 acgccctgga cgccgtggtg gtggcctgca gcaccgtggc catgcagcag aagatcaccc
     6721 gcttcgtgcg ctacaaggag atgaacgcct tcgacggtaa aaccatcgac aaggagaccg
     6781 gcgaggtgct gcaccagaag acccacttcc cccagccctg ggagttcttc gcccaggagg
     6841 tgatgatccg cgtgttcggc aagcccgacg gcaagcccga gttcgaggag gccgacaccc
     6901 ccgagaagct gcgcaccctg ctggccgaga agctgagcag ccgccctgag gccgtgcacg
     6961 agtacgtgac tcctctgttc gtgagccgcg cccccaaccg caagatgagc ggtcagggtc
     7021 acatggagac cgtgaagagc gccaagcgcc tggacgaggg cgtgagcgtg ctgcgcgtgc
     7081 ccctgaccca gctgaagctg aaggacctgg agaagatggt gaaccgcgag cgcgagccca
     7141 agctgtacga ggccctgaag gcccgcctgg aggcccacaa ggacgacccc gccaaggcct
     7201 tcgccgagcc cttctacaag tacgacaagg ccggcaaccg cacccagcag gtgaaggccg
     7261 tgcgcgtgga gcaggtgcag aagaccggcg tgtgggtgcg caaccacaac ggcatcgccg
     7321 acaacgccac catggtgcgc gtggacgtgt tcgagaaggg cgacaagtac tacctggtgc
     7381 ccatctacag ctggcaggtg gccaagggca tcctgcccga ccgcgccgtg gtgcagggca
     7441 aggacgagga ggactggcag ctgatcgacg acagcttcaa cttcaagttc agcctgcacc
     7501 ccaacgacct ggtggaggtg atcaccaaga aggcccgcat gttcggctac ttcgccagct
     7561 gccaccgcgg caccggcaac atcaacatcc gcatccacga cctggaccac aagatcggca
     7621 agaacggcat cctggagggc atcggcgtga agaccgccct gagcttccag aagtaccaga
     7681 tcgacgagct gggcaaggag atccgcccct gccgcctgaa gaagcgccct cctgtgcgca
     7741 gcagggctga ccccaagaag aagaggaagg tggaggccag cctcgagaaa aggccggcgg
     7801 ccacgaaaaa ggccggccag gcaaaaaaga aaaagggcag ttaatgatct agtaatgatc
     7861 tagaactata gtgagtcgta ttacgtagat ccagacatga taagatacat tgatgagttt
     7921 ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct
     7981 attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt
     8041 cattttatgt ttcaggttca gggggaggtg tgggaggttt tttaattcgc ggccgcggcg
     8101 ccaatgcatt gggcccggta cccagctttt gttcccttta gtgagggtta attgcgcgct
     8161 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     8221 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     8281 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     8341 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     8401 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     8461 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     8521 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     8581 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     8641 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     8701 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     8761 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     8821 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     8881 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     8941 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     9001 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     9061 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     9121 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     9181 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     9241 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     9301 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     9361 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     9421 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     9481 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     9541 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     9601 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     9661 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     9721 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     9781 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     9841 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     9901 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     9961 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
    10021 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
    10081 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
    10141 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
    10201 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
    10261 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
    10321 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
    10381 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
    10441 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
    10501 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
    10561 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
    10621 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
    10681 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
    10741 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
    10801 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agtcgaccat
    10861 agccaattca atatggcgta tatggactca tgccaattca atatggtgga tctggacctg
    10921 tgccaattca atatggcgta tatggactcg tgccaattca atatggtgga tctggacccc
    10981 agccaattca atatggcgga cttggcacca tgccaattca atatggcgga cttggcactg
    11041 tgccaactgg ggaggggtct acttggcacg gtgccaagtt tgaggagggg tcttggccct
    11101 gtgccaagtc cgccatattg aattggcatg gtgccaataa tggcggccat attggctata
    11161 tgccaggatc aatatatagg caatatccaa tatggcccta tgccaatatg gctattggcc
    11221 aggttcaata ctatgtattg gccctatgcc atatagtatt ccatatatgg gttttcctat
    11281 tgacgtagat agcccctccc aatgggcggt cccatatacc atatatgggg cttcctaata
    11341 ccgcccatag ccactccccc attgacgtca atggtctcta tatatggtct ttcctattga
    11401 cgtcatatgg gcggtcctat tgacgtatat ggcgcctccc ccattgacgt caattacggt
    11461 aaatggcccg cctggctcaa tgcccattga cgtcaatagg accacccacc attgacgtca
    11521 atgggatggc tcattgccca ttcatatccg ttctcacgcc ccctattgac gtcaatgacg
    11581 gtaaatggcc cacttggcag tacatcaata tctattaata gtaacttggc aagtacatta
    11641 ctattggaag gacgccaggg tacattggca gtactcccat tgacgtcaat ggcggtaaat
    11701 ggcccgcgat ggctgccaag tacatcccca ttgacgtcaa tggggagggg caatgacgca
    11761 aatgggcgtt ccattgacgt aaatgggcgg taggcgtgcc taatgggagg tctatataag
    11821 caatgctcgt ttaggga
//
