LOCUS       sequence-122268-        8465 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses wild type SpCas9 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8465)
  AUTHORS   Bolukbasi MF, Gupta A, Oikemus S, Derr AG, Garber M, Brodsky MH, Zhu
            LJ, Wolfe SA
  TITLE     DNA-binding-domain fusions enhance the targeting range and precision
            of Cas9.
  JOURNAL   Nat Methods. 2015 Oct 19. doi: 10.1038/nmeth.3624.
  PUBMED    26480473
REFERENCE   2  (bases 1 to 8465)
  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 
            Methods. 2015 Oct 19. doi: 10.1038/nmeth.3624."
COMMENT     Sequence Label: pCSDest2-SpCas9-NLS-3XHA-NLS
FEATURES             Location/Qualifiers
     source          1..8465
                     /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"
     protein_bind    110..134
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      138..147
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             144..4247
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             144..4247
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             144..4247
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             4260..4280
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4281..4370
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     CDS             4383..4409
                     /codon_start=1
                     /product="nuclear localization signal (Makkerh et al., 
                     1996)"
                     /label=NLS
                     /translation="PAAKKKKLD"
     protein_bind    complement(4447..4471)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    4516..4650
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4540..4559)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4594..4613
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4763..4783)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4763..4781)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4802..4818)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4802..4818)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4815..4837)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4826..4842
                     /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(4850..4856,4857..4874,4875..4880))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4895..4916
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5033..5050)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5204..5792)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5284..5303)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5963..6754,6755..6823))
                     /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     6586..6605
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6824..6928)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6954..7409)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7091..7112)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7303..7322
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(7483..8465,1..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 caaggatcaa caagtttgta
      121 caaaaaagca ggctggcgcc accatggaca agaagtactc cattgggctc gatatcggta
      181 ccaacagcgt cggctgggcc gtcattacgg acgagtacaa ggtgccgagc aaaaaattca
      241 aagttctggg caataccgat cgccacagca taaagaagaa cctcattgga gccctcctgt
      301 tcgactccgg ggagacggcc gaagccacgc ggctcaaaag aacagcacgg cgcagatata
      361 cccgcagaaa gaatcggatc tgctacctgc aggagatctt tagtaatgag atggctaagg
      421 tggatgactc tttcttccat aggctggagg agtccttttt ggtggaggag gataaaaagc
      481 acgagcgcca cccaatcttt ggcaatatcg tggacgaggt ggcgtaccat gaaaagtacc
      541 caaccatata tcatctgagg aagaagctgg tagacagtac tgataaggct gacttgcggt
      601 tgatctatct cgcgctggcg cacatgatca aatttcgggg acacttcctc atcgaggggg
      661 acctgaaccc agacaacagc gatgtcgaca aactctttat ccaactggtt cagacttaca
      721 atcagctttt cgaggagaac ccgatcaacg catccggcgt tgacgccaaa gcaatcctga
      781 gcgctaggct gtccaaatcc cggcggctcg aaaacctcat cgcacagctc cctggggaga
      841 agaagaacgg cctgtttggt aatcttatcg ccctgtcact cgggctgacc cccaacttta
      901 aatctaactt cgacctggcc gaagatgcca agctgcaact gagcaaagac acctacgatg
      961 atgatctcga caatctgctg gcccagatcg gcgaccagta cgcagacctt tttttggcgg
     1021 caaagaacct gtcagacgcc attctgctga gtgatattct gcgagtgaac acggagatca
     1081 ccaaagctcc gctgagcgct agtatgatca agcgctatga tgagcaccac caagacttga
     1141 ctttgctgaa ggcccttgtc agacagcaac tgcctgagaa gtacaaggaa attttcttcg
     1201 atcagtctaa aaatggctac gccggataca ttgacggcgg agcaagccag gaggaatttt
     1261 acaaatttat taagcccatc ttggaaaaaa tggacggcac cgaggagctg ctggtaaagc
     1321 tgaacagaga agatctgttg cgcaaacagc gcactttcga caatggaagc atcccccacc
     1381 agattcacct gggcgaactg cacgctatcc tcaggcggca agaggatttc tacccctttt
     1441 tgaaagataa cagggaaaag attgagaaaa tcctcacatt tcggataccc tactatgtag
     1501 gccccctcgc tcggggaaat tccagattcg cgtggatgac tcgcaaatca gaagagacca
     1561 tcactccctg gaacttcgag gaagtcgtgg ataagggggc ctctgcccag tccttcatcg
     1621 aaaggatgac taactttgat aaaaatctgc ctaacgaaaa ggtgcttcct aaacactctc
     1681 tgctgtacga gtacttcaca gtttataacg agctcaccaa ggtcaaatac gtcacagaag
     1741 ggatgagaaa gccagcattc ctgtctggag agcagaagaa agctatcgtg gacctcctct
     1801 tcaagacgaa ccggaaagtt accgtgaaac agctcaaaga agactatttc aaaaagattg
     1861 aatgtttcga ctctgttgaa atcagcggag tggaggatcg cttcaacgca tccctgggaa
     1921 cgtatcacga tctcctgaaa atcattaaag acaaggactt cctggacaat gaggagaacg
     1981 aggacattct tgaggacatt gtcctcaccc ttacgttgtt tgaagatagg gagatgattg
     2041 aagaacgctt gaaaacttac gctcatctct tcgacgacaa agtcatgaaa cagctcaaga
     2101 gacgccgata tacaggatgg gggcggctgt caagaaaact gatcaatggc atccgagaca
     2161 agcagagtgg aaagacaatc ctggattttc ttaagtccga tggatttgcc aaccggaact
     2221 tcatgcagtt gatccatgat gactctctca cctttaagga ggacatccag aaagcacaag
     2281 tttctggcca gggggacagt cttcacgagc acatcgctaa tcttgcaggt agcccagcta
     2341 tcaaaaaggg aatactgcag accgttaagg tcgtggatga actcgtcaaa gtaatgggaa
     2401 ggcataagcc cgagaatatc gttatcgaga tggcccgaga gaaccaaact acccagaagg
     2461 gacagaagaa cagtagggaa aggatgaaga ggattgaaga gggtataaaa gaactggggt
     2521 cccaaatcct taaggaacac ccagttgaaa acacccagct tcagaatgag aagctctacc
     2581 tgtactacct gcagaacggc agggacatgt acgtggatca ggaactggac atcaaccggt
     2641 tgtccgacta cgacgtggat catatcgtgc cccaaagctt tctcaaagat gattctattg
     2701 ataataaagt gttgacaaga tccgataaaa atagagggaa gagtgataac gtcccctcag
     2761 aagaagttgt caagaaaatg aaaaattatt ggcggcagct gctgaacgcc aaactgatca
     2821 cacaacggaa gttcgataat ctgactaagg ctgaacgagg tggcctgtct gagttggata
     2881 aagccggctt catcaaaagg cagcttgttg agacacgcca gatcaccaag cacgtggccc
     2941 aaattctcga ttcacgcatg aacaccaagt acgatgaaaa tgacaaactg attcgagagg
     3001 tgaaagttat tactctgaag tctaagctgg tctcagattt cagaaaggac tttcagtttt
     3061 ataaggtgag agagatcaac aattaccacc atgcgcatga tgcctacctg aatgcagtgg
     3121 taggcactgc acttatcaaa aaatatccca agctggaatc tgaatttgtt tacggagact
     3181 ataaagtgta cgatgttagg aaaatgatcg caaagtctga gcaggaaata ggcaaggcca
     3241 ccgctaagta cttcttttac agcaatatta tgaatttttt caagaccgag attacactgg
     3301 ccaatggaga gattcggaag cgaccactta tcgaaacaaa cggagaaaca ggagaaatcg
     3361 tgtgggacaa gggtagggat ttcgcgacag tccgcaaggt cctgtccatg ccgcaggtga
     3421 acatcgttaa aaagaccgaa gtacagaccg gaggcttctc caaggaaagt atcctcccga
     3481 aaaggaacag cgacaagctg atcgcacgca aaaaagattg ggaccccaag aaatacggcg
     3541 gattcgattc tcctacagtc gcttacagtg tactggttgt ggccaaagtg gagaaaggga
     3601 agtctaaaaa actcaaaagc gtcaaggaac tgctgggcat cacaatcatg gagcgatcca
     3661 gcttcgagaa aaaccccatc gactttctcg aagcgaaagg atataaagag gtcaaaaaag
     3721 acctcatcat taagctgccc aagtactctc tctttgagct tgaaaacggc cggaaacgaa
     3781 tgctcgctag tgcgggcgag ctgcagaaag gtaacgagct ggcactgccc tctaaatacg
     3841 ttaatttctt gtatctggcc agccactatg aaaagctcaa agggtctccc gaagataatg
     3901 agcagaagca gctgttcgtg gaacaacaca aacactacct tgatgagatc atcgagcaaa
     3961 taagcgagtt ctccaaaaga gtgatcctcg ccgacgctaa cctcgataag gtgctttctg
     4021 cttacaataa gcacagggat aagcccatca gggagcaggc agaaaacatt atccacttgt
     4081 ttactctgac caacttgggc gcgcctgcag ccttcaagta cttcgacacc accatagaca
     4141 gaaagcggta cacctctaca aaggaggtcc tggacgccac actgattcat cagtcaatta
     4201 cggggctcta tgaaacaaga atcgacctct ctcagctcgg tggagacggc accggcgggc
     4261 ccaagaagaa gaggaaggta tacccatacg atgttcctga ctatgcgggc tatccctatg
     4321 acgtcccgga ctatgcagga tcgtatcctt atgacgttcc agattacgct ggatccgccg
     4381 ctccggcagc taagaaaaag aaactggatt tcgaatccgg ataagcggcc gcctcgagat
     4441 atctagaccc agctttcttg tacaaagtgg ttgatcctct cgagcctcta gaactatagt
     4501 gagtcgtatt acgtagatcc agacatgata agatacattg atgagtttgg acaaaccaca
     4561 actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt
     4621 gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt
     4681 caggttcagg gggaggtgtg ggaggttttt taattcgcgg ccgcggcgcc aatgcattgg
     4741 gcccggtacc cagcttttgt tccctttagt gagggttaat tgcgcgcttg gcgtaatcat
     4801 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     4861 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     4921 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4981 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     5041 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     5101 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     5161 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     5221 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     5281 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     5341 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     5401 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     5461 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     5521 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     5581 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     5641 gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     5701 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     5761 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     5821 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     5881 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     5941 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     6001 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     6061 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     6121 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     6181 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     6241 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     6301 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     6361 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     6421 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     6481 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     6541 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     6601 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     6661 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     6721 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     6781 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     6841 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     6901 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctaaattgt
     6961 aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa
     7021 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt
     7081 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa
     7141 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag
     7201 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
     7261 tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
     7321 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
     7381 cgcgcttaat gcgccgctac agggcgcgtc ccattcgcca ttcaggctgc gcaactgttg
     7441 ggaagggcga tcggtgcggg cctcttcgct attacgccag tcgaccatag ccaattcaat
     7501 atggcgtata tggactcatg ccaattcaat atggtggatc tggacctgtg ccaattcaat
     7561 atggcgtata tggactcgtg ccaattcaat atggtggatc tggaccccag ccaattcaat
     7621 atggcggact tggcaccatg ccaattcaat atggcggact tggcactgtg ccaactgggg
     7681 aggggtctac ttggcacggt gccaagtttg aggaggggtc ttggccctgt gccaagtccg
     7741 ccatattgaa ttggcatggt gccaataatg gcggccatat tggctatatg ccaggatcaa
     7801 tatataggca atatccaata tggccctatg ccaatatggc tattggccag gttcaatact
     7861 atgtattggc cctatgccat atagtattcc atatatgggt tttcctattg acgtagatag
     7921 cccctcccaa tgggcggtcc catataccat atatggggct tcctaatacc gcccatagcc
     7981 actcccccat tgacgtcaat ggtctctata tatggtcttt cctattgacg tcatatgggc
     8041 ggtcctattg acgtatatgg cgcctccccc attgacgtca attacggtaa atggcccgcc
     8101 tggctcaatg cccattgacg tcaataggac cacccaccat tgacgtcaat gggatggctc
     8161 attgcccatt catatccgtt ctcacgcccc ctattgacgt caatgacggt aaatggccca
     8221 cttggcagta catcaatatc tattaatagt aacttggcaa gtacattact attggaagga
     8281 cgccagggta cattggcagt actcccattg acgtcaatgg cggtaaatgg cccgcgatgg
     8341 ctgccaagta catccccatt gacgtcaatg gggaggggca atgacgcaaa tgggcgttcc
     8401 attgacgtaa atgggcggta ggcgtgccta atgggaggtc tatataagca atgctcgttt
     8461 aggga
//
