LOCUS       sequence-177716-       10976 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of modified SpyCas9 in e.coli..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10976)
  AUTHORS   Staahl BT, Benekareddy M, Coulon-Bainier C, Banfal AA, Floor SN, 
            Sabo JK, Urnes C, Munares GA, Ghosh A, Doudna JA
  TITLE     Efficient genome editing in the mouse brain by local delivery of 
            engineered Cas9 ribonucleoprotein complexes.
  JOURNAL   Nat Biotechnol. 2017 Feb 13. doi: 10.1038/nbt.3806.
  PUBMED    28191903
REFERENCE   2  (bases 1 to 10976)
  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 
            Biotechnol. 2017 Feb 13. doi: 10.1038/nbt.3806."
COMMENT     Sequence Label: 4xNLS-pMJ915v2-sfGFP
FEATURES             Location/Qualifiers
     source          1..10976
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             3718..3738
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3745..3765
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4614..4632)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4628..4675
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(5041..5046,5047..5063,5064..5069))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(5096..5114)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5182..5286
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5287..5355,5356..6147)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5505..5524)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6318..6906
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6807..6826
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7060..7077
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    7092..7231
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(7157..7176)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7317..7339
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(7333..7524)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     8982..9001
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     9085..9104
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        9085..9103
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             9192..9214
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             9234..9251
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             9261..10361
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     10335..10358
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             10416..10436
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             10446..10466
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             10476..10496
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             10506..10526
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             10536..10556
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(10584..10976,1..3711)
                     /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             join(10584..10976,1..3711)
                     /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"
ORIGIN
        1 tatccaacta tctatcatct gcgaaaaaaa ttggtagatt ctactgataa agcggatttg
       61 cgcttaatct atttggcctt agcgcatatg attaagtttc gtggtcattt tttgattgag
      121 ggagatttaa atcctgataa tagtgatgtg gacaaactat ttatccagtt ggtacaaacc
      181 tacaatcaat tatttgaaga aaaccctatt aacgcaagtg gagtagatgc taaagcgatt
      241 ctttctgcac gattgagtaa atcaagacga ttagaaaatc tcattgctca gctccccggt
      301 gagaagaaaa atggcttatt tgggaatctc attgctttgt cattgggttt gacccctaat
      361 tttaaatcaa attttgattt ggcagaagat gctaaattac agctttcaaa agatacttac
      421 gatgatgatt tagataattt attggcgcaa attggagatc aatatgctga tttgtttttg
      481 gcagctaaga atttatcaga tgctatttta ctttcagata tcctaagagt aaatactgaa
      541 ataactaagg ctcccctatc agcttcaatg attaaacgct acgatgaaca tcatcaagac
      601 ttgactcttt taaaagcttt agttcgacaa caacttccag aaaagtataa agaaatcttt
      661 tttgatcaat caaaaaacgg atatgcaggt tatattgatg ggggagctag ccaagaagaa
      721 ttttataaat ttatcaaacc aattttagaa aaaatggatg gtactgagga attattggtg
      781 aaactaaatc gtgaagattt gctgcgcaag caacggacct ttgacaacgg ctctattccc
      841 catcaaattc acttgggtga gctgcatgct attttgagaa gacaagaaga cttttatcca
      901 tttttaaaag acaatcgtga gaagattgaa aaaatcttga cttttcgaat tccttattat
      961 gttggtccat tggcgcgtgg caatagtcgt tttgcatgga tgactcggaa gtctgaagaa
     1021 acaattaccc catggaattt tgaagaagtt gtcgataaag gtgcttcagc tcaatcattt
     1081 attgaacgca tgacaaactt tgataaaaat cttccaaatg aaaaagtact accaaaacat
     1141 agtttgcttt atgagtattt tacggtttat aacgaattga caaaggtcaa atatgttact
     1201 gaaggaatgc gaaaaccagc atttctttca ggtgaacaga agaaagccat tgttgattta
     1261 ctcttcaaaa caaatcgaaa agtaaccgtt aagcaattaa aagaagatta tttcaaaaaa
     1321 atagaatgtt ttgatagtgt tgaaatttca ggagttgaag atagatttaa tgcttcatta
     1381 ggtacctacc atgatttgct aaaaattatt aaagataaag attttttgga taatgaagaa
     1441 aatgaagata tcttagagga tattgtttta acattgacct tatttgaaga tagggagatg
     1501 attgaggaaa gacttaaaac atatgctcac ctctttgatg ataaggtgat gaaacagctt
     1561 aaacgtcgcc gttatactgg ttggggacgt ttgtctcgaa aattgattaa tggtattagg
     1621 gataagcaat ctggcaaaac aatattagat tttttgaaat cagatggttt tgccaatcgc
     1681 aattttatgc agctgatcca tgatgatagt ttgacattta aagaagacat tcaaaaagca
     1741 caagtgtctg gacaaggcga tagtttacat gaacatattg caaatttagc tggtagccct
     1801 gctattaaaa aaggtatttt acagactgta aaagttgttg atgaattggt caaagtaatg
     1861 gggcggcata agccagaaaa tatcgttatt gaaatggcac gtgaaaatca gacaactcaa
     1921 aagggccaga aaaattcgcg agagcgtatg aaacgaatcg aagaaggtat caaagaatta
     1981 ggaagtcaga ttcttaaaga gcatcctgtt gaaaatactc aattgcaaaa tgaaaagctc
     2041 tatctctatt atctccaaaa tggaagagac atgtatgtgg accaagaatt agatattaat
     2101 cgtttaagtg attatgatgt cgatcacatt gttccacaaa gtttccttaa agacgattca
     2161 atagacaata aggtcttaac gcgttctgat aaaaatcgtg gtaaatcgga taacgttcca
     2221 agtgaagaag tagtcaaaaa gatgaaaaac tattggagac aacttctaaa cgccaagtta
     2281 atcactcaac gtaagtttga taatttaacg aaagctgaac gtggaggttt gagtgaactt
     2341 gataaagctg gttttatcaa acgccaattg gttgaaactc gccaaatcac taagcatgtg
     2401 gcacaaattt tggatagtcg catgaatact aaatacgatg aaaatgataa acttattcga
     2461 gaggttaaag tgattacctt aaaatctaaa ttagtttctg acttccgaaa agatttccaa
     2521 ttctataaag tacgtgagat taacaattac catcatgccc atgatgcgta tctaaatgcc
     2581 gtcgttggaa ctgctttgat taagaaatat ccaaaacttg aatcggagtt tgtctatggt
     2641 gattataaag tttatgatgt tcgtaaaatg attgctaagt ctgagcaaga aataggcaaa
     2701 gcaaccgcaa aatatttctt ttactctaat atcatgaact tcttcaaaac agaaattaca
     2761 cttgcaaatg gagagattcg caaacgccct ctaatcgaaa ctaatgggga aactggagaa
     2821 attgtctggg ataaagggcg agattttgcc acagtgcgca aagtattgtc catgccccaa
     2881 gtcaatattg tcaagaaaac agaagtacag acaggcggat tctccaagga gtcaatttta
     2941 ccaaaaagaa attcggacaa gcttattgct cgtaaaaaag actgggatcc aaaaaaatat
     3001 ggtggttttg atagtccaac ggtagcttat tcagtcctag tggttgctaa ggtggaaaaa
     3061 gggaaatcga agaagttaaa atccgttaaa gagttactag ggatcacaat tatggaaaga
     3121 agttcctttg aaaaaaatcc gattgacttt ttagaagcta aaggatataa ggaagttaaa
     3181 aaagacttaa tcattaaact acctaaatat agtctttttg agttagaaaa cggtcgtaaa
     3241 cggatgctgg ctagtgccgg agaattacaa aaaggaaatg agctggctct gccaagcaaa
     3301 tatgtgaatt ttttatattt agctagtcat tatgaaaagt tgaagggtag tccagaagat
     3361 aacgaacaaa aacaattgtt tgtggagcag cataagcatt atttagatga gattattgag
     3421 caaatcagtg aattttctaa gcgtgttatt ttagcagatg ccaatttaga taaagttctt
     3481 agtgcatata acaaacatag agacaaacca atacgtgaac aagcagaaaa tattattcat
     3541 ttatttacgt tgacgaatct tggagctccc gctgctttta aatattttga tacaacaatt
     3601 gatcgtaaac gatatacgtc tacaaaagaa gttttagatg ccactcttat ccatcaatcc
     3661 atcactggtc tttatgaaac acgcattgat ttgagtcagc taggaggtga cgggtcacct
     3721 aagaaaaaac gaaaagttga ggatcctaaa aagaaacgaa aagttgatac cggttctaaa
     3781 ggtgaagaac tgttcaccgg tgttgttccg atcctggttg aactggatgg tgatgttaac
     3841 ggccacaaat tctctgttcg tggtgaaggt gaaggtgatg caaccaacgg taaactgacc
     3901 ctgaaattca tctgcactac cggtaaactg ccggttccat ggccgactct ggtgactacc
     3961 ctgacctatg gtgttcagtg tttttctcgt tacccggatc acatgaagca gcatgatttc
     4021 ttcaaatctg caatgccgga aggttatgta caggagcgca ccatttcttt caaagacgat
     4081 ggcacctaca aaacccgtgc agaggttaaa tttgaaggtg atactctggt gaaccgtatt
     4141 gaactgaaag gcattgattt caaagaggac ggcaacatcc tgggccacaa actggaatat
     4201 aacttcaact cccataacgt ttacatcacc gcagacaaac agaagaacgg tatcaaagct
     4261 aacttcaaaa ttcgccataa cgttgaagac ggtagcgtac agctggcgga ccactaccag
     4321 cagaacactc cgatcggtga tggtccggtt ctgctgccgg ataaccacta cctgtccacc
     4381 cagtctgttc tgtccaaaga cccgaacgaa aagcgcgacc acatggtgct gctggagttc
     4441 gttactgcag caggtatcac gcacggcatg gatgagctct acaaataata acattggaag
     4501 tggataacgg atccgcgatc gcggcgcgcc acctggtggc cggccggtac cacgcgtgcg
     4561 cgctgatccg gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga
     4621 gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa
     4681 aggaggaact atatccggat atccacagga cgggtgtggt cgccatgatc gcgtagtcga
     4741 tagtggctcc aagtagcgaa gcgagcagga ctgggcggcg gccaaagcgg tcggacagtg
     4801 ctccgagaac gggtgcgcat agaaattgca tcaacgcata tagcgctagc agcacgccat
     4861 agtgactggc gatgctgtcg gaatggacga tatcccgcaa gaggcccggc agtaccggca
     4921 taaccaagcc tatgcctaca gcatccaggg tgacggtgcc gaggatgacg atgagcgcat
     4981 tgttagattt catacacggt gcctgactgc gttagcaatt taactgtgat aaactaccgc
     5041 attaaagctt atcgatgata agctgtcaaa catgagaatt cttgaagacg aaagggcctc
     5101 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     5161 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     5221 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataaca ttgaaaaagg
     5281 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     5341 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     5401 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     5461 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     5521 ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     5581 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     5641 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     5701 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     5761 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     5821 acgatgcctg cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     5881 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     5941 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     6001 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     6061 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     6121 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     6181 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     6241 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     6301 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     6361 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     6421 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg
     6481 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     6541 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     6601 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     6661 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     6721 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     6781 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     6841 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     6901 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     6961 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     7021 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     7081 gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     7141 aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc
     7201 gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc
     7261 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     7321 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta
     7381 aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag
     7441 ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag
     7501 ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg
     7561 ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca
     7621 tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca
     7681 gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca
     7741 gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt
     7801 ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt
     7861 cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg
     7921 ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat
     7981 gcgcacccgt ggccaggacc caacgctgcc cgagatgcgc cgcgtgcggc tgctggagat
     8041 ggcggacgcg atggatatgt tctgccaagg gttggtttgc gcattcacag ttctccgcaa
     8101 gaattgattg gctccaattc ttggagtggt gaatccgtta gcgaggtgcc gccggcttcc
     8161 attcaggtcg aggtggcccg gctccatgca ccgcgacgca acgcggggag gcagacaagg
     8221 tatagggcgg cgcctacaat ccatgccaac ccgttccatg tgctcgccga ggcggcataa
     8281 atcgccgtga cgatcagcgg tccaatgatc gaagttaggc tggtaagagc cgcgagcgat
     8341 ccttgaagct gtccctgatg gtcgtcatct acctgcctgg acagcatggc ctgcaacgcg
     8401 ggcatcccga tgccgccgga agcgagaaga atcataatgg ggaaggccat ccagcctcgc
     8461 gtcgcgaacg ccagcaagac gtagcccagc gcgtcggccg ccatgccggc gataatggcc
     8521 tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc gagggcgtgc
     8581 aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc
     8641 tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat gataaagaag
     8701 acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga gctgactggg
     8761 ttgaaggctc tcaagggcat cggtcgacgc tctcccttat gcgactcctg cattaggaag
     8821 cagcccagta gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag
     8881 gagatggcgc ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa
     8941 gcgctcatga gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag
     9001 gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg
     9061 atcgagatct cgatcccgcg aaattaatac gactcactat agggagacca caacggtttc
     9121 cctctagtgc cggctccgga gagctcttta attaagcggc cgccctgcag gactcgagtt
     9181 ctagaaataa ttttgtttaa ctttaagaag gagatataca tatgaaatct tctcaccatc
     9241 accatcacca tggttcttct atgaaaatcg aagaaggtaa actggtaatc tggattaacg
     9301 gcgataaagg ctataacggt ctcgctgaag tcggtaagaa attcgagaaa gataccggaa
     9361 ttaaagtcac cgttgagcat ccggataaac tggaagagaa attcccacag gttgcggcaa
     9421 ctggcgatgg ccctgacatt atcttctggg cacacgaccg ctttggtggc tacgctcaat
     9481 ctggcctgtt ggctgaaatc accccggaca aagcgttcca ggacaagctg tatccgttta
     9541 cctgggatgc cgtacgttac aacggcaagc tgattgctta cccgatcgct gttgaagcgt
     9601 tatcgctgat ttataacaaa gatctgctgc cgaacccgcc aaaaacctgg gaagagatcc
     9661 cggcgctgga taaagaactg aaagcgaaag gtaagagcgc gctgatgttc aacctgcaag
     9721 aaccgtactt cacctggccg ctgattgctg ctgacggggg ttatgcgttc aagtatgaaa
     9781 acggcaagta cgacattaaa gacgtgggcg tggataacgc tggcgcgaaa gcgggtctga
     9841 ccttcctggt tgacctgatt aaaaacaaac acatgaatgc agacaccgat tactccatcg
     9901 cagaagctgc ctttaataaa ggcgaaacag cgatgaccat caacggcccg tgggcatggt
     9961 ccaacatcga caccagcaaa gtgaattatg gtgtaacggt actgccgacc ttcaagggtc
    10021 aaccatccaa accgttcgtt ggcgtgctga gcgcaggtat taacgccgcc agtccgaaca
    10081 aagagctggc aaaagagttc ctcgaaaact atctgctgac tgatgaaggt ctggaagcgg
    10141 ttaataaaga caaaccgctg ggtgccgtag cgctgaagtc ttacgaggaa gagttggcga
    10201 aagatccacg tattgccgcc actatggaaa acgcccagaa aggtgaaatc atgccgaaca
    10261 tcccgcagat gtccgctttc tggtatgccg tgcgtactgc ggtgatcaac gccgccagcg
    10321 gtcgtcagac tgtcgatgaa gccctgaaag acgcgcagac taattcgagc tcgaacaaca
    10381 acaacaataa caataacaac aacctcggga tcgaggaaaa cctgtacttc caatccaatg
    10441 ccacccccaa gaagaagcga aaggtgggtg ggtccccaaa gaagaaacgg aaagtaggcg
    10501 gctcccccaa aaagaagcga aaagtagggg gtagccccaa gaagaagcgg aaggtaggta
    10561 tccacggagt cccagcagct accatggata agaaatactc aataggctta gatatcggca
    10621 caaatagcgt cggatgggcg gtgatcactg atgaatataa ggttccgtct aaaaagttca
    10681 aggttctggg aaatacagac cgccacagta tcaaaaaaaa tcttataggg gctcttttat
    10741 ttgacagtgg agagacagcg gaagcgactc gtctcaaacg gacagctcgt agaaggtata
    10801 cacgtcggaa gaatcgtatt tgttatctac aggagatttt ttcaaatgag atggcgaaag
    10861 tagatgatag tttctttcat cgacttgaag agtctttttt ggtggaagaa gacaagaagc
    10921 atgaacgtca tcctattttt ggaaatatag tagatgaagt tgcttatcat gagaaa
//
