LOCUS       sequence-177419-       10886 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 10886)
  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 10886)
  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: 1xNLS-pMJ915v2-sfGFP
FEATURES             Location/Qualifiers
     source          1..10886
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1034..1057
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1115..1135
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1145..1165
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1193..5296
                     /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             1193..5296
                     /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             5303..5323
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5330..5350
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(6199..6217)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6213..6260
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(6626..6631,6632..6648,6649..6654))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(6681..6699)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6767..6871
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6872..6940,6941..7732)
                     /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(7090..7109)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7903..8491
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8392..8411
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8645..8662
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    8677..8816
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(8742..8761)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8902..8924
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(8918..9109)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     10567..10586
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     10670..10689
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        10670..10688
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             10777..10799
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             10819..10836
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             join(10846..10886,1..1060)
                     /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"
ORIGIN
        1 cgataaaggc tataacggtc tcgctgaagt cggtaagaaa ttcgagaaag ataccggaat
       61 taaagtcacc gttgagcatc cggataaact ggaagagaaa ttcccacagg ttgcggcaac
      121 tggcgatggc cctgacatta tcttctgggc acacgaccgc tttggtggct acgctcaatc
      181 tggcctgttg gctgaaatca ccccggacaa agcgttccag gacaagctgt atccgtttac
      241 ctgggatgcc gtacgttaca acggcaagct gattgcttac ccgatcgctg ttgaagcgtt
      301 atcgctgatt tataacaaag atctgctgcc gaacccgcca aaaacctggg aagagatccc
      361 ggcgctggat aaagaactga aagcgaaagg taagagcgcg ctgatgttca acctgcaaga
      421 accgtacttc acctggccgc tgattgctgc tgacgggggt tatgcgttca agtatgaaaa
      481 cggcaagtac gacattaaag acgtgggcgt ggataacgct ggcgcgaaag cgggtctgac
      541 cttcctggtt gacctgatta aaaacaaaca catgaatgca gacaccgatt actccatcgc
      601 agaagctgcc tttaataaag gcgaaacagc gatgaccatc aacggcccgt gggcatggtc
      661 caacatcgac accagcaaag tgaattatgg tgtaacggta ctgccgacct tcaagggtca
      721 accatccaaa ccgttcgttg gcgtgctgag cgcaggtatt aacgccgcca gtccgaacaa
      781 agagctggca aaagagttcc tcgaaaacta tctgctgact gatgaaggtc tggaagcggt
      841 taataaagac aaaccgctgg gtgccgtagc gctgaagtct tacgaggaag agttggcgaa
      901 agatccacgt attgccgcca ctatggaaaa cgcccagaaa ggtgaaatca tgccgaacat
      961 cccgcagatg tccgctttct ggtatgccgt gcgtactgcg gtgatcaacg ccgccagcgg
     1021 tcgtcagact gtcgatgaag ccctgaaaga cgcgcagact aattcgagct cgaacaacaa
     1081 caacaataac aataacaaca acctcgggat cgaggaaaac ctgtacttcc aatccaatgc
     1141 caccccaaag aagaaacgga aagtaggcat ccacggagtc ccagcagcta ccatggataa
     1201 gaaatactca ataggcttag atatcggcac aaatagcgtc ggatgggcgg tgatcactga
     1261 tgaatataag gttccgtcta aaaagttcaa ggttctggga aatacagacc gccacagtat
     1321 caaaaaaaat cttatagggg ctcttttatt tgacagtgga gagacagcgg aagcgactcg
     1381 tctcaaacgg acagctcgta gaaggtatac acgtcggaag aatcgtattt gttatctaca
     1441 ggagattttt tcaaatgaga tggcgaaagt agatgatagt ttctttcatc gacttgaaga
     1501 gtcttttttg gtggaagaag acaagaagca tgaacgtcat cctatttttg gaaatatagt
     1561 agatgaagtt gcttatcatg agaaatatcc aactatctat catctgcgaa aaaaattggt
     1621 agattctact gataaagcgg atttgcgctt aatctatttg gccttagcgc atatgattaa
     1681 gtttcgtggt cattttttga ttgagggaga tttaaatcct gataatagtg atgtggacaa
     1741 actatttatc cagttggtac aaacctacaa tcaattattt gaagaaaacc ctattaacgc
     1801 aagtggagta gatgctaaag cgattctttc tgcacgattg agtaaatcaa gacgattaga
     1861 aaatctcatt gctcagctcc ccggtgagaa gaaaaatggc ttatttggga atctcattgc
     1921 tttgtcattg ggtttgaccc ctaattttaa atcaaatttt gatttggcag aagatgctaa
     1981 attacagctt tcaaaagata cttacgatga tgatttagat aatttattgg cgcaaattgg
     2041 agatcaatat gctgatttgt ttttggcagc taagaattta tcagatgcta ttttactttc
     2101 agatatccta agagtaaata ctgaaataac taaggctccc ctatcagctt caatgattaa
     2161 acgctacgat gaacatcatc aagacttgac tcttttaaaa gctttagttc gacaacaact
     2221 tccagaaaag tataaagaaa tcttttttga tcaatcaaaa aacggatatg caggttatat
     2281 tgatggggga gctagccaag aagaatttta taaatttatc aaaccaattt tagaaaaaat
     2341 ggatggtact gaggaattat tggtgaaact aaatcgtgaa gatttgctgc gcaagcaacg
     2401 gacctttgac aacggctcta ttccccatca aattcacttg ggtgagctgc atgctatttt
     2461 gagaagacaa gaagactttt atccattttt aaaagacaat cgtgagaaga ttgaaaaaat
     2521 cttgactttt cgaattcctt attatgttgg tccattggcg cgtggcaata gtcgttttgc
     2581 atggatgact cggaagtctg aagaaacaat taccccatgg aattttgaag aagttgtcga
     2641 taaaggtgct tcagctcaat catttattga acgcatgaca aactttgata aaaatcttcc
     2701 aaatgaaaaa gtactaccaa aacatagttt gctttatgag tattttacgg tttataacga
     2761 attgacaaag gtcaaatatg ttactgaagg aatgcgaaaa ccagcatttc tttcaggtga
     2821 acagaagaaa gccattgttg atttactctt caaaacaaat cgaaaagtaa ccgttaagca
     2881 attaaaagaa gattatttca aaaaaataga atgttttgat agtgttgaaa tttcaggagt
     2941 tgaagataga tttaatgctt cattaggtac ctaccatgat ttgctaaaaa ttattaaaga
     3001 taaagatttt ttggataatg aagaaaatga agatatctta gaggatattg ttttaacatt
     3061 gaccttattt gaagataggg agatgattga ggaaagactt aaaacatatg ctcacctctt
     3121 tgatgataag gtgatgaaac agcttaaacg tcgccgttat actggttggg gacgtttgtc
     3181 tcgaaaattg attaatggta ttagggataa gcaatctggc aaaacaatat tagatttttt
     3241 gaaatcagat ggttttgcca atcgcaattt tatgcagctg atccatgatg atagtttgac
     3301 atttaaagaa gacattcaaa aagcacaagt gtctggacaa ggcgatagtt tacatgaaca
     3361 tattgcaaat ttagctggta gccctgctat taaaaaaggt attttacaga ctgtaaaagt
     3421 tgttgatgaa ttggtcaaag taatggggcg gcataagcca gaaaatatcg ttattgaaat
     3481 ggcacgtgaa aatcagacaa ctcaaaaggg ccagaaaaat tcgcgagagc gtatgaaacg
     3541 aatcgaagaa ggtatcaaag aattaggaag tcagattctt aaagagcatc ctgttgaaaa
     3601 tactcaattg caaaatgaaa agctctatct ctattatctc caaaatggaa gagacatgta
     3661 tgtggaccaa gaattagata ttaatcgttt aagtgattat gatgtcgatc acattgttcc
     3721 acaaagtttc cttaaagacg attcaataga caataaggtc ttaacgcgtt ctgataaaaa
     3781 tcgtggtaaa tcggataacg ttccaagtga agaagtagtc aaaaagatga aaaactattg
     3841 gagacaactt ctaaacgcca agttaatcac tcaacgtaag tttgataatt taacgaaagc
     3901 tgaacgtgga ggtttgagtg aacttgataa agctggtttt atcaaacgcc aattggttga
     3961 aactcgccaa atcactaagc atgtggcaca aattttggat agtcgcatga atactaaata
     4021 cgatgaaaat gataaactta ttcgagaggt taaagtgatt accttaaaat ctaaattagt
     4081 ttctgacttc cgaaaagatt tccaattcta taaagtacgt gagattaaca attaccatca
     4141 tgcccatgat gcgtatctaa atgccgtcgt tggaactgct ttgattaaga aatatccaaa
     4201 acttgaatcg gagtttgtct atggtgatta taaagtttat gatgttcgta aaatgattgc
     4261 taagtctgag caagaaatag gcaaagcaac cgcaaaatat ttcttttact ctaatatcat
     4321 gaacttcttc aaaacagaaa ttacacttgc aaatggagag attcgcaaac gccctctaat
     4381 cgaaactaat ggggaaactg gagaaattgt ctgggataaa gggcgagatt ttgccacagt
     4441 gcgcaaagta ttgtccatgc cccaagtcaa tattgtcaag aaaacagaag tacagacagg
     4501 cggattctcc aaggagtcaa ttttaccaaa aagaaattcg gacaagctta ttgctcgtaa
     4561 aaaagactgg gatccaaaaa aatatggtgg ttttgatagt ccaacggtag cttattcagt
     4621 cctagtggtt gctaaggtgg aaaaagggaa atcgaagaag ttaaaatccg ttaaagagtt
     4681 actagggatc acaattatgg aaagaagttc ctttgaaaaa aatccgattg actttttaga
     4741 agctaaagga tataaggaag ttaaaaaaga cttaatcatt aaactaccta aatatagtct
     4801 ttttgagtta gaaaacggtc gtaaacggat gctggctagt gccggagaat tacaaaaagg
     4861 aaatgagctg gctctgccaa gcaaatatgt gaatttttta tatttagcta gtcattatga
     4921 aaagttgaag ggtagtccag aagataacga acaaaaacaa ttgtttgtgg agcagcataa
     4981 gcattattta gatgagatta ttgagcaaat cagtgaattt tctaagcgtg ttattttagc
     5041 agatgccaat ttagataaag ttcttagtgc atataacaaa catagagaca aaccaatacg
     5101 tgaacaagca gaaaatatta ttcatttatt tacgttgacg aatcttggag ctcccgctgc
     5161 ttttaaatat tttgatacaa caattgatcg taaacgatat acgtctacaa aagaagtttt
     5221 agatgccact cttatccatc aatccatcac tggtctttat gaaacacgca ttgatttgag
     5281 tcagctagga ggtgacgggt cacctaagaa aaaacgaaaa gttgaggatc ctaaaaagaa
     5341 acgaaaagtt gataccggtt ctaaaggtga agaactgttc accggtgttg ttccgatcct
     5401 ggttgaactg gatggtgatg ttaacggcca caaattctct gttcgtggtg aaggtgaagg
     5461 tgatgcaacc aacggtaaac tgaccctgaa attcatctgc actaccggta aactgccggt
     5521 tccatggccg actctggtga ctaccctgac ctatggtgtt cagtgttttt ctcgttaccc
     5581 ggatcacatg aagcagcatg atttcttcaa atctgcaatg ccggaaggtt atgtacagga
     5641 gcgcaccatt tctttcaaag acgatggcac ctacaaaacc cgtgcagagg ttaaatttga
     5701 aggtgatact ctggtgaacc gtattgaact gaaaggcatt gatttcaaag aggacggcaa
     5761 catcctgggc cacaaactgg aatataactt caactcccat aacgtttaca tcaccgcaga
     5821 caaacagaag aacggtatca aagctaactt caaaattcgc cataacgttg aagacggtag
     5881 cgtacagctg gcggaccact accagcagaa cactccgatc ggtgatggtc cggttctgct
     5941 gccggataac cactacctgt ccacccagtc tgttctgtcc aaagacccga acgaaaagcg
     6001 cgaccacatg gtgctgctgg agttcgttac tgcagcaggt atcacgcacg gcatggatga
     6061 gctctacaaa taataacatt ggaagtggat aacggatccg cgatcgcggc gcgccacctg
     6121 gtggccggcc ggtaccacgc gtgcgcgctg atccggctgc taacaaagcc cgaaaggaag
     6181 ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac
     6241 gggtcttgag gggttttttg ctgaaaggag gaactatatc cggatatcca caggacgggt
     6301 gtggtcgcca tgatcgcgta gtcgatagtg gctccaagta gcgaagcgag caggactggg
     6361 cggcggccaa agcggtcgga cagtgctccg agaacgggtg cgcatagaaa ttgcatcaac
     6421 gcatatagcg ctagcagcac gccatagtga ctggcgatgc tgtcggaatg gacgatatcc
     6481 cgcaagaggc ccggcagtac cggcataacc aagcctatgc ctacagcatc cagggtgacg
     6541 gtgccgagga tgacgatgag cgcattgtta gatttcatac acggtgcctg actgcgttag
     6601 caatttaact gtgataaact accgcattaa agcttatcga tgataagctg tcaaacatga
     6661 gaattcttga agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     6721 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     6781 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     6841 aatgcttcaa taacattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     6901 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     6961 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     7021 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     7081 taaagttctg ctatgtggcg cggtattatc ccgtgttgac gccgggcaag agcaactcgg
     7141 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     7201 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     7261 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     7321 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     7381 cataccaaac gacgagcgtg acaccacgat gcctgcagca atggcaacaa cgttgcgcaa
     7441 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     7501 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     7561 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     7621 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     7681 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     7741 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     7801 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     7861 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     7921 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     7981 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     8041 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     8101 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     8161 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     8221 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     8281 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     8341 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     8401 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     8461 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     8521 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     8581 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     8641 gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac
     8701 gcatctgtgc ggtatttcac accgcaatgg tgcactctca gtacaatctg ctctgatgcc
     8761 gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc
     8821 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     8881 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     8941 cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga
     9001 tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc
     9061 ttctgataaa gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg
     9121 tgtaaggggg atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca
     9181 cgatacgggt tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac
     9241 tggcggtatg gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg
     9301 ttaatacaga tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga
     9361 acataatggt gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga
     9421 agaccattca tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc
     9481 gctcgcgtat cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg
     9541 tcctcaacga caggagcacg atcatgcgca cccgtggcca ggacccaacg ctgcccgaga
     9601 tgcgccgcgt gcggctgctg gagatggcgg acgcgatgga tatgttctgc caagggttgg
     9661 tttgcgcatt cacagttctc cgcaagaatt gattggctcc aattcttgga gtggtgaatc
     9721 cgttagcgag gtgccgccgg cttccattca ggtcgaggtg gcccggctcc atgcaccgcg
     9781 acgcaacgcg gggaggcaga caaggtatag ggcggcgcct acaatccatg ccaacccgtt
     9841 ccatgtgctc gccgaggcgg cataaatcgc cgtgacgatc agcggtccaa tgatcgaagt
     9901 taggctggta agagccgcga gcgatccttg aagctgtccc tgatggtcgt catctacctg
     9961 cctggacagc atggcctgca acgcgggcat cccgatgccg ccggaagcga gaagaatcat
    10021 aatggggaag gccatccagc ctcgcgtcgc gaacgccagc aagacgtagc ccagcgcgtc
    10081 ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt
    10141 gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat
    10201 cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg
    10261 tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg
    10321 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gacgctctcc
    10381 cttatgcgac tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc
    10441 cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg ccacggggcc
    10501 tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag cccgatcttc
    10561 cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc
    10621 ggccacgatg cgtccggcgt agaggatcga gatctcgatc ccgcgaaatt aatacgactc
    10681 actataggga gaccacaacg gtttccctct agtgccggct ccggagagct ctttaattaa
    10741 gcggccgccc tgcaggactc gagttctaga aataattttg tttaacttta agaaggagat
    10801 atacatatga aatcttctca ccatcaccat caccatggtt cttctatgaa aatcgaagaa
    10861 ggtaaactgg taatctggat taacgg
//
