LOCUS       sequence-204775-       11834 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 11834)
  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 11834)
  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..11834
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        34..52
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     34..51
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             102..149
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             168..188
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             195..4295
                     /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             195..4295
                     /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             195..4295
                     /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             4308..4328
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4329..4418
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     CDS             4431..4457
                     /codon_start=1
                     /product="nuclear localization signal (Makkerh et al., 
                     1996)"
                     /label=NLS
                     /translation="PAAKKKKLD"
     CDS             4473..4493
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(4494..4535,4536..4538,4539..6248,6249..6254,6255..6320,
                     6321..6323,6324..7736)
                     /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      7323..7332
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             7749..7769
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7785..7832
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     polyA_signal    7885..8019
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7909..7928)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     7963..7982
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(8132..8152)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(8132..8150)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(8171..8187)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8171..8187)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(8184..8206)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    8195..8211
                     /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(8219..8225,8226..8243,8244..8249))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8264..8285
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8402..8419)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8573..9161)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8653..8672)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9332..10123,10124..10192))
                     /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     9955..9974
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(10193..10297)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(10323..10778)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10460..10481)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     10672..10691
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        10849..11833
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 gccattctgc ctggggacgt cggagcaagc ttgatttagg tgacactata gaatacaagc
       61 tacttgttct ttttgcagga tcccatcgat tcgaattcat gaaaaggccg gcggccacga
      121 aaaaggccgg ccaggcaaaa aagaaaaagc tggacggcag cggaagcccc aagaagaaga
      181 ggaaggtacc atgggacaag aagtactcca ttgggctcga tatcggtacc aacagcgtcg
      241 gctgggccgt cattacggac gagtacaagg tgccgagcaa aaaattcaaa gttctgggca
      301 ataccgatcg ccacagcata aagaagaacc tcattggagc cctcctgttc gactccgggg
      361 agacggccga agccacgcgg ctcaaaagaa cagcacggcg cagatatacc cgcagaaaga
      421 atcggatctg ctacctgcag gagatcttta gtaatgagat ggctaaggtg gatgactctt
      481 tcttccatag gctggaggag tcctttttgg tggaggagga taaaaagcac gagcgccacc
      541 caatctttgg caatatcgtg gacgaggtgg cgtaccatga aaagtaccca accatatatc
      601 atctgaggaa gaagctggta gacagtactg ataaggctga cttgcggttg atctatctcg
      661 cgctggcgca catgatcaaa tttcggggac acttcctcat cgagggggac ctgaacccag
      721 acaacagcga tgtcgacaaa ctctttatcc aactggttca gacttacaat cagcttttcg
      781 aggagaaccc gatcaacgca tccggcgttg acgccaaagc aatcctgagc gctaggctgt
      841 ccaaatcccg gcggctcgaa aacctcatcg cacagctccc tggggagaag aagaacggcc
      901 tgtttggtaa tcttatcgcc ctgtcactcg ggctgacccc caactttaaa tctaacttcg
      961 acctggccga agatgccaag ctgcaactga gcaaagacac ctacgatgat gatctcgaca
     1021 atctgctggc ccagatcggc gaccagtacg cagacctttt tttggcggca aagaacctgt
     1081 cagacgccat tctgctgagt gatattctgc gagtgaacac ggagatcacc aaagctccgc
     1141 tgagcgctag tatgatcaag cgctatgatg agcaccacca agacttgact ttgctgaagg
     1201 cccttgtcag acagcaactg cctgagaagt acaaggaaat tttcttcgat cagtctaaaa
     1261 atggctacgc cggatacatt gacggcggag caagccagga ggaattttac aaatttatta
     1321 agcccatctt ggaaaaaatg gacggcaccg aggagctgct ggtaaagctg aacagagaag
     1381 atctgttgcg caaacagcgc actttcgaca atggaagcat cccccaccag attcacctgg
     1441 gcgaactgca cgctatcctc aggcggcaag aggatttcta cccctttttg aaagataaca
     1501 gggaaaagat tgagaaaatc ctcacatttc ggatacccta ctatgtaggc cccctcgctc
     1561 ggggaaattc cagattcgcg tggatgactc gcaaatcaga agagaccatc actccctgga
     1621 acttcgagga agtcgtggat aagggggcct ctgcccagtc cttcatcgaa aggatgacta
     1681 actttgataa aaatctgcct aacgaaaagg tgcttcctaa acactctctg ctgtacgagt
     1741 acttcacagt ttataacgag ctcaccaagg tcaaatacgt cacagaaggg atgagaaagc
     1801 cagcattcct gtctggagag cagaagaaag ctatcgtgga cctcctcttc aagacgaacc
     1861 ggaaagttac cgtgaaacag ctcaaagaag actatttcaa aaagattgaa tgtttcgact
     1921 ctgttgaaat cagcggagtg gaggatcgct tcaacgcatc cctgggaacg tatcacgatc
     1981 tcctgaaaat cattaaagac aaggacttcc tggacaatga ggagaacgag gacattcttg
     2041 aggacattgt cctcaccctt acgttgtttg aagataggga gatgattgaa gaacgcttga
     2101 aaacttacgc tcatctcttc gacgacaaag tcatgaaaca gctcaagaga cgccgatata
     2161 caggatgggg gcggctgtca agaaaactga tcaatggcat ccgagacaag cagagtggaa
     2221 agacaatcct ggattttctt aagtccgatg gatttgccaa ccggaacttc atgcagttga
     2281 tccatgatga ctctctcacc tttaaggagg acatccagaa agcacaagtt tctggccagg
     2341 gggacagtct tcacgagcac atcgctaatc ttgcaggtag cccagctatc aaaaagggaa
     2401 tactgcagac cgttaaggtc gtggatgaac tcgtcaaagt aatgggaagg cataagcccg
     2461 agaatatcgt tatcgagatg gcccgagaga accaaactac ccagaaggga cagaagaaca
     2521 gtagggaaag gatgaagagg attgaagagg gtataaaaga actggggtcc caaatcctta
     2581 aggaacaccc agttgaaaac acccagcttc agaatgagaa gctctacctg tactacctgc
     2641 agaacggcag ggacatgtac gtggatcagg aactggacat caaccggttg tccgactacg
     2701 acgtggatca tatcgtgccc caaagctttc tcaaagatga ttctattgat aataaagtgt
     2761 tgacaagatc cgataaaaat agagggaaga gtgataacgt cccctcagaa gaagttgtca
     2821 agaaaatgaa aaattattgg cggcagctgc tgaacgccaa actgatcaca caacggaagt
     2881 tcgataatct gactaaggct gaacgaggtg gcctgtctga gttggataaa gccggcttca
     2941 tcaaaaggca gcttgttgag acacgccaga tcaccaagca cgtggcccaa attctcgatt
     3001 cacgcatgaa caccaagtac gatgaaaatg acaaactgat tcgagaggtg aaagttatta
     3061 ctctgaagtc taagctggtc tcagatttca gaaaggactt tcagttttat aaggtgagag
     3121 agatcaacaa ttaccaccat gcgcatgatg cctacctgaa tgcagtggta ggcactgcac
     3181 ttatcaaaaa atatcccaag ctggaatctg aatttgttta cggagactat aaagtgtacg
     3241 atgttaggaa aatgatcgca aagtctgagc aggaaatagg caaggccacc gctaagtact
     3301 tcttttacag caatattatg aattttttca agaccgagat tacactggcc aatggagaga
     3361 ttcggaagcg accacttatc gaaacaaacg gagaaacagg agaaatcgtg tgggacaagg
     3421 gtagggattt cgcgacagtc cgcaaggtcc tgtccatgcc gcaggtgaac atcgttaaaa
     3481 agaccgaagt acagaccgga ggcttctcca aggaaagtat cctcccgaaa aggaacagcg
     3541 acaagctgat cgcacgcaaa aaagattggg accccaagaa atacggcgga ttcgattctc
     3601 ctacagtcgc ttacagtgta ctggttgtgg ccaaagtgga gaaagggaag tctaaaaaac
     3661 tcaaaagcgt caaggaactg ctgggcatca caatcatgga gcgatccagc ttcgagaaaa
     3721 accccatcga ctttctcgaa gcgaaaggat ataaagaggt caaaaaagac ctcatcatta
     3781 agctgcccaa gtactctctc tttgagcttg aaaacggccg gaaacgaatg ctcgctagtg
     3841 cgggcgagct gcagaaaggt aacgagctgg cactgccctc taaatacgtt aatttcttgt
     3901 atctggccag ccactatgaa aagctcaaag ggtctcccga agataatgag cagaagcagc
     3961 tgttcgtgga acaacacaaa cactaccttg atgagatcat cgagcaaata agcgagttct
     4021 ccaaaagagt gatcctcgcc gacgctaacc tcgataaggt gctttctgct tacaataagc
     4081 acagggataa gcccatcagg gagcaggcag aaaacattat ccacttgttt actctgacca
     4141 acttgggcgc gcctgcagcc ttcaagtact tcgacaccac catagacaga aagcggtaca
     4201 cctctacaaa ggaggtcctg gacgccacac tgattcatca gtcaattacg gggctctatg
     4261 aaacaagaat cgacctctct cagctcggtg gagacggcac cggcgggccc aagaagaaga
     4321 ggaaggtata cccatacgat gttcctgact atgcgggcta tccctatgac gtcccggact
     4381 atgcaggatc gtatccttat gacgttccag attacgctgg atccgccgct ccggcagcta
     4441 agaaaaagaa actggatttc gaatccggaa tgcccaagaa gaagaggaag gtggccgcct
     4501 tcaagcccaa ccccatcaac tacatcctgg gcctggccat cggcatcgcc agcgtgggct
     4561 gggccatggt ggagatcgac gaggacgaga accccatctg cctgatcgac ctgggtgtgc
     4621 gcgtgttcga gcgcgctgag gtgcccaaga ctggtgacag tctggctatg gctcgccggc
     4681 ttgctcgctc tgttcggcgc cttactcgcc ggcgcgctca ccgccttctg cgcgctcgcc
     4741 gcctgctgaa gcgcgagggt gtgctgcagg ctgccgactt cgacgagaac ggcctgatca
     4801 agagcctgcc caacactcct tggcagctgc gcgctgccgc tctggaccgc aagctgactc
     4861 ctctggagtg gagcgccgtg ctgctgcacc tgatcaagca ccgcggctac ctgagccagc
     4921 gcaagaacga gggcgagacc gccgacaagg agctgggtgc tctgctgaag ggcgtggccg
     4981 acaacgccca cgccctgcag actggtgact tccgcactcc tgctgagctg gccctgaaca
     5041 agttcgagaa ggagagcggc cacatccgca accagcgcgg cgactacagc cacaccttca
     5101 gccgcaagga cctgcaggcc gagctgatcc tgctgttcga gaagcagaag gagttcggca
     5161 acccccacgt gagcggcggc ctgaaggagg gcatcgagac cctgctgatg acccagcgcc
     5221 ccgccctgag cggcgacgcc gtgcagaaga tgctgggcca ctgcaccttc gagccagccg
     5281 agcccaaggc cgccaagaac acctacaccg ccgagcgctt catctggctg accaagctga
     5341 acaacctgcg catcctggag cagggcagcg agcgccccct gaccgacacc gagcgcgcca
     5401 ccctgatgga cgagccctac cgcaagagca agctgaccta cgcccaggcc cgcaagctgc
     5461 tgggtctgga ggacaccgcc ttcttcaagg gcctgcgcta cggcaaggac aacgccgagg
     5521 ccagcaccct gatggagatg aaggcctacc acgccatcag ccgcgccctg gagaaggagg
     5581 gcctgaagga caagaagagt cctctgaacc tgagccccga gctgcaggac gagatcggca
     5641 ccgccttcag cctgttcaag accgacgagg acatcaccgg ccgcctgaag gaccgcatcc
     5701 agcccgagat cctggaggcc ctgctgaagc acatcagctt cgacaagttc gtgcagatca
     5761 gcctgaaggc cctgcgccgc atcgtgcccc tgatggagca gggcaagcgc tacgacgagg
     5821 cctgcgccga gatctacggc gaccactacg gcaagaagaa caccgaggag aagatctacc
     5881 tgcctcctat ccccgccgac gagatccgca accccgtggt gctgcgcgcc ctgagccagg
     5941 cccgcaaggt gatcaacggc gtggtgcgcc gctacggcag ccccgcccgc atccacatcg
     6001 agaccgcccg cgaggtgggc aagagcttca aggaccgcaa ggagatcgag aagcgccagg
     6061 aggagaaccg caaggaccgc gagaaggccg ccgccaagtt ccgcgagtac ttccccaact
     6121 tcgtgggcga gcccaagagc aaggacatcc tgaagctgcg cctgtacgag cagcagcacg
     6181 gcaagtgcct gtacagcggc aaggagatca acctgggccg cctgaacgag aagggctacg
     6241 tggagatcgc cgctgccctg cccttcagcc gcacctggga cgacagcttc aacaacaagg
     6301 tgctggtgct gggcagcgag gctcagaaca agggcaacca gaccccctac gagtacttca
     6361 acggcaagga caacagccgc gagtggcagg agttcaaggc ccgcgtggag accagccgct
     6421 tcccccgcag caagaagcag cgcatcctgc tgcagaagtt cgacgaggac ggcttcaagg
     6481 agcgcaacct gaacgacacc cgctacgtga accgcttcct gtgccagttc gtggccgacc
     6541 gcatgcgcct gaccggcaag ggcaagaagc gcgtgttcgc cagcaacggc cagatcacca
     6601 acctgctgcg cggcttctgg ggcctgcgca aggtgcgcgc cgagaacgac cgccaccacg
     6661 ccctggacgc cgtggtggtg gcctgcagca ccgtggccat gcagcagaag atcacccgct
     6721 tcgtgcgcta caaggagatg aacgccttcg acggtaaaac catcgacaag gagaccggcg
     6781 aggtgctgca ccagaagacc cacttccccc agccctggga gttcttcgcc caggaggtga
     6841 tgatccgcgt gttcggcaag cccgacggca agcccgagtt cgaggaggcc gacacccccg
     6901 agaagctgcg caccctgctg gccgagaagc tgagcagccg ccctgaggcc gtgcacgagt
     6961 acgtgactcc tctgttcgtg agccgcgccc ccaaccgcaa gatgagcggt cagggtcaca
     7021 tggagaccgt gaagagcgcc aagcgcctgg acgagggcgt gagcgtgctg cgcgtgcccc
     7081 tgacccagct gaagctgaag gacctggaga agatggtgaa ccgcgagcgc gagcccaagc
     7141 tgtacgaggc cctgaaggcc cgcctggagg cccacaagga cgaccccgcc aaggccttcg
     7201 ccgagccctt ctacaagtac gacaaggccg gcaaccgcac ccagcaggtg aaggccgtgc
     7261 gcgtggagca ggtgcagaag accggcgtgt gggtgcgcaa ccacaacggc atcgccgaca
     7321 acgccaccat ggtgcgcgtg gacgtgttcg agaagggcga caagtactac ctggtgccca
     7381 tctacagctg gcaggtggcc aagggcatcc tgcccgaccg cgccgtggtg cagggcaagg
     7441 acgaggagga ctggcagctg atcgacgaca gcttcaactt caagttcagc ctgcacccca
     7501 acgacctggt ggaggtgatc accaagaagg cccgcatgtt cggctacttc gccagctgcc
     7561 accgcggcac cggcaacatc aacatccgca tccacgacct ggaccacaag atcggcaaga
     7621 acggcatcct ggagggcatc ggcgtgaaga ccgccctgag cttccagaag taccagatcg
     7681 acgagctggg caaggagatc cgcccctgcc gcctgaagaa gcgccctcct gtgcgcagca
     7741 gggctgaccc caagaagaag aggaaggtgg aggccagcct cgagaaaagg ccggcggcca
     7801 cgaaaaaggc cggccaggca aaaaagaaaa agggcagtta atgatctagt aatgatctag
     7861 aactatagtg agtcgtatta cgtagatcca gacatgataa gatacattga tgagtttgga
     7921 caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt
     7981 gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa ttgcattcat
     8041 tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt aattcgcggc cgcggcgcca
     8101 atgcattggg cccggtaccc agcttttgtt ccctttagtg agggttaatt gcgcgcttgg
     8161 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     8221 acatacgagc cgggagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     8281 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     8341 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     8401 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     8461 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     8521 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     8581 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     8641 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     8701 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     8761 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     8821 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     8881 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     8941 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     9001 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     9061 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     9121 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     9181 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     9241 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     9301 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     9361 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     9421 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     9481 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     9541 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     9601 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     9661 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     9721 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     9781 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     9841 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     9901 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     9961 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
    10021 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
    10081 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
    10141 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
    10201 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
    10261 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
    10321 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
    10381 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
    10441 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
    10501 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
    10561 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
    10621 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
    10681 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
    10741 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
    10801 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagt cgaccgccaa
    10861 ttcaatatgg cgtatatgga ctcatgccaa ttcaatatgg tggatctgga cctgtgccaa
    10921 ttcaatatgg cgtatatgga ctcgtgccaa ttcaatatgg tggatctgga ccccagccaa
    10981 ttcaatatgg cggacttggc accatgccaa ttcaatatgg cggacctggc actgtgccaa
    11041 ctggggaggg gtctacttgg cacggtgcca agtttgagga ggggtcttgg ccctgtgcca
    11101 agtccgccat attgaattgg catggtgcca ataatggcgg ccatattggc tatatgccag
    11161 gatcaatata taggcaatat ccaatatggc cctatgccaa tatggctatt ggccaggttc
    11221 aatactatgt attggcccta tgccatatag tattccatat atgggttttc ctattgacgt
    11281 agatagcccc tcccaatggg cggtcccata taccatatat ggggcttcct aataccgccc
    11341 atagccactc ccccattgac gtcaatggtc tctatatatg gtctttccta ttgacgtcat
    11401 atgggcggtc ctattgacgt atatggcgcc tcccccattg acgtcaatta cggtaaatgg
    11461 cccgcctggc tcaatgccca ttgacgtcaa taggaccacc caccattgac gtcaatggga
    11521 tggctcattg cccattcata tccgttctca cgccccctat tgacgtcaat gacggtaaat
    11581 ggcccacttg gcagtacatc aatatctatt aatagtaact tggcaagtac attactattg
    11641 gaagtacgcc agggtacatt ggcagtactc ccattgacgt caatggcggt aaatggcccg
    11701 cgatggctgc caagtacatc cccattgacg tcaatgggga ggggcaatga cgcaaatggg
    11761 cgttccattg acgtaaatgg gcggtaggcg tgcctaatgg gaggtctata taagcaatgc
    11821 tcgtttaggg aacc
//
