LOCUS       sequence-177325-       10133 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 10133)
  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 10133)
  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: pMJ915v2+Nterm Spe1 +Cterm Age1 site
FEATURES             Location/Qualifiers
     source          1..10133
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     97..116
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     350..367
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    382..521
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(447..466)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     607..629
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(623..814)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     2272..2291
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2375..2394
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2375..2393
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             2482..2504
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2524..2541
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2551..3651
                     /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     3625..3648
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             3706..3726
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             3742..7845
                     /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             3742..7845
                     /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             7852..7872
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7879..7899
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(8037..8055)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      8051..8098
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(8464..8469,8470..8486,8487..8492))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(8519..8537)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        8605..8709
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8710..8778,8779..9570)
                     /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(8928..8947)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(9741..10133,1..196)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
       61 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
      121 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
      181 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
      241 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
      301 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
      361 gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
      421 ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
      481 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
      541 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
      601 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
      661 agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat
      721 ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg
      781 ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc
      841 tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg
      901 atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc
      961 ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag
     1021 gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg
     1081 gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg
     1141 ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg
     1201 attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga
     1261 gcacgatcat gcgcacccgt ggccaggacc caacgctgcc cgagatgcgc cgcgtgcggc
     1321 tgctggagat ggcggacgcg atggatatgt tctgccaagg gttggtttgc gcattcacag
     1381 ttctccgcaa gaattgattg gctccaattc ttggagtggt gaatccgtta gcgaggtgcc
     1441 gccggcttcc attcaggtcg aggtggcccg gctccatgca ccgcgacgca acgcggggag
     1501 gcagacaagg tatagggcgg cgcctacaat ccatgccaac ccgttccatg tgctcgccga
     1561 ggcggcataa atcgccgtga cgatcagcgg tccaatgatc gaagttaggc tggtaagagc
     1621 cgcgagcgat ccttgaagct gtccctgatg gtcgtcatct acctgcctgg acagcatggc
     1681 ctgcaacgcg ggcatcccga tgccgccgga agcgagaaga atcataatgg ggaaggccat
     1741 ccagcctcgc gtcgcgaacg ccagcaagac gtagcccagc gcgtcggccg ccatgccggc
     1801 gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc
     1861 gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg
     1921 aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat
     1981 gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga
     2041 gctgactggg ttgaaggctc tcaagggcat cggtcgacgc tctcccttat gcgactcctg
     2101 cattaggaag cagcccagta gtaggttgag gccgttgagc accgccgccg caaggaatgg
     2161 tgcatgcaag gagatggcgc ccaacagtcc cccggccacg gggcctgcca ccatacccac
     2221 gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga tcttccccat cggtgatgtc
     2281 ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc
     2341 ggcgtagagg atcgagatct cgatcccgcg aaattaatac gactcactat agggagacca
     2401 caacggtttc cctctagtgc cggctccgga gagctcttta attaagcggc cgccctgcag
     2461 gactcgagtt ctagaaataa ttttgtttaa ctttaagaag gagatataca tatgaaatct
     2521 tctcaccatc accatcacca tggttcttct atgaaaatcg aagaaggtaa actggtaatc
     2581 tggattaacg gcgataaagg ctataacggt ctcgctgaag tcggtaagaa attcgagaaa
     2641 gataccggaa ttaaagtcac cgttgagcat ccggataaac tggaagagaa attcccacag
     2701 gttgcggcaa ctggcgatgg ccctgacatt atcttctggg cacacgaccg ctttggtggc
     2761 tacgctcaat ctggcctgtt ggctgaaatc accccggaca aagcgttcca ggacaagctg
     2821 tatccgttta cctgggatgc cgtacgttac aacggcaagc tgattgctta cccgatcgct
     2881 gttgaagcgt tatcgctgat ttataacaaa gatctgctgc cgaacccgcc aaaaacctgg
     2941 gaagagatcc cggcgctgga taaagaactg aaagcgaaag gtaagagcgc gctgatgttc
     3001 aacctgcaag aaccgtactt cacctggccg ctgattgctg ctgacggggg ttatgcgttc
     3061 aagtatgaaa acggcaagta cgacattaaa gacgtgggcg tggataacgc tggcgcgaaa
     3121 gcgggtctga ccttcctggt tgacctgatt aaaaacaaac acatgaatgc agacaccgat
     3181 tactccatcg cagaagctgc ctttaataaa ggcgaaacag cgatgaccat caacggcccg
     3241 tgggcatggt ccaacatcga caccagcaaa gtgaattatg gtgtaacggt actgccgacc
     3301 ttcaagggtc aaccatccaa accgttcgtt ggcgtgctga gcgcaggtat taacgccgcc
     3361 agtccgaaca aagagctggc aaaagagttc ctcgaaaact atctgctgac tgatgaaggt
     3421 ctggaagcgg ttaataaaga caaaccgctg ggtgccgtag cgctgaagtc ttacgaggaa
     3481 gagttggcga aagatccacg tattgccgcc actatggaaa acgcccagaa aggtgaaatc
     3541 atgccgaaca tcccgcagat gtccgctttc tggtatgccg tgcgtactgc ggtgatcaac
     3601 gccgccagcg gtcgtcagac tgtcgatgaa gccctgaaag acgcgcagac taattcgagc
     3661 tcgaacaaca acaacaataa caataacaac aacctcggga tcgaggaaaa cctgtacttc
     3721 caatccaatg ccactagtac catggataag aaatactcaa taggcttaga tatcggcaca
     3781 aatagcgtcg gatgggcggt gatcactgat gaatataagg ttccgtctaa aaagttcaag
     3841 gttctgggaa atacagaccg ccacagtatc aaaaaaaatc ttataggggc tcttttattt
     3901 gacagtggag agacagcgga agcgactcgt ctcaaacgga cagctcgtag aaggtataca
     3961 cgtcggaaga atcgtatttg ttatctacag gagatttttt caaatgagat ggcgaaagta
     4021 gatgatagtt tctttcatcg acttgaagag tcttttttgg tggaagaaga caagaagcat
     4081 gaacgtcatc ctatttttgg aaatatagta gatgaagttg cttatcatga gaaatatcca
     4141 actatctatc atctgcgaaa aaaattggta gattctactg ataaagcgga tttgcgctta
     4201 atctatttgg ccttagcgca tatgattaag tttcgtggtc attttttgat tgagggagat
     4261 ttaaatcctg ataatagtga tgtggacaaa ctatttatcc agttggtaca aacctacaat
     4321 caattatttg aagaaaaccc tattaacgca agtggagtag atgctaaagc gattctttct
     4381 gcacgattga gtaaatcaag acgattagaa aatctcattg ctcagctccc cggtgagaag
     4441 aaaaatggct tatttgggaa tctcattgct ttgtcattgg gtttgacccc taattttaaa
     4501 tcaaattttg atttggcaga agatgctaaa ttacagcttt caaaagatac ttacgatgat
     4561 gatttagata atttattggc gcaaattgga gatcaatatg ctgatttgtt tttggcagct
     4621 aagaatttat cagatgctat tttactttca gatatcctaa gagtaaatac tgaaataact
     4681 aaggctcccc tatcagcttc aatgattaaa cgctacgatg aacatcatca agacttgact
     4741 cttttaaaag ctttagttcg acaacaactt ccagaaaagt ataaagaaat cttttttgat
     4801 caatcaaaaa acggatatgc aggttatatt gatgggggag ctagccaaga agaattttat
     4861 aaatttatca aaccaatttt agaaaaaatg gatggtactg aggaattatt ggtgaaacta
     4921 aatcgtgaag atttgctgcg caagcaacgg acctttgaca acggctctat tccccatcaa
     4981 attcacttgg gtgagctgca tgctattttg agaagacaag aagactttta tccattttta
     5041 aaagacaatc gtgagaagat tgaaaaaatc ttgacttttc gaattcctta ttatgttggt
     5101 ccattggcgc gtggcaatag tcgttttgca tggatgactc ggaagtctga agaaacaatt
     5161 accccatgga attttgaaga agttgtcgat aaaggtgctt cagctcaatc atttattgaa
     5221 cgcatgacaa actttgataa aaatcttcca aatgaaaaag tactaccaaa acatagtttg
     5281 ctttatgagt attttacggt ttataacgaa ttgacaaagg tcaaatatgt tactgaagga
     5341 atgcgaaaac cagcatttct ttcaggtgaa cagaagaaag ccattgttga tttactcttc
     5401 aaaacaaatc gaaaagtaac cgttaagcaa ttaaaagaag attatttcaa aaaaatagaa
     5461 tgttttgata gtgttgaaat ttcaggagtt gaagatagat ttaatgcttc attaggtacc
     5521 taccatgatt tgctaaaaat tattaaagat aaagattttt tggataatga agaaaatgaa
     5581 gatatcttag aggatattgt tttaacattg accttatttg aagataggga gatgattgag
     5641 gaaagactta aaacatatgc tcacctcttt gatgataagg tgatgaaaca gcttaaacgt
     5701 cgccgttata ctggttgggg acgtttgtct cgaaaattga ttaatggtat tagggataag
     5761 caatctggca aaacaatatt agattttttg aaatcagatg gttttgccaa tcgcaatttt
     5821 atgcagctga tccatgatga tagtttgaca tttaaagaag acattcaaaa agcacaagtg
     5881 tctggacaag gcgatagttt acatgaacat attgcaaatt tagctggtag ccctgctatt
     5941 aaaaaaggta ttttacagac tgtaaaagtt gttgatgaat tggtcaaagt aatggggcgg
     6001 cataagccag aaaatatcgt tattgaaatg gcacgtgaaa atcagacaac tcaaaagggc
     6061 cagaaaaatt cgcgagagcg tatgaaacga atcgaagaag gtatcaaaga attaggaagt
     6121 cagattctta aagagcatcc tgttgaaaat actcaattgc aaaatgaaaa gctctatctc
     6181 tattatctcc aaaatggaag agacatgtat gtggaccaag aattagatat taatcgttta
     6241 agtgattatg atgtcgatca cattgttcca caaagtttcc ttaaagacga ttcaatagac
     6301 aataaggtct taacgcgttc tgataaaaat cgtggtaaat cggataacgt tccaagtgaa
     6361 gaagtagtca aaaagatgaa aaactattgg agacaacttc taaacgccaa gttaatcact
     6421 caacgtaagt ttgataattt aacgaaagct gaacgtggag gtttgagtga acttgataaa
     6481 gctggtttta tcaaacgcca attggttgaa actcgccaaa tcactaagca tgtggcacaa
     6541 attttggata gtcgcatgaa tactaaatac gatgaaaatg ataaacttat tcgagaggtt
     6601 aaagtgatta ccttaaaatc taaattagtt tctgacttcc gaaaagattt ccaattctat
     6661 aaagtacgtg agattaacaa ttaccatcat gcccatgatg cgtatctaaa tgccgtcgtt
     6721 ggaactgctt tgattaagaa atatccaaaa cttgaatcgg agtttgtcta tggtgattat
     6781 aaagtttatg atgttcgtaa aatgattgct aagtctgagc aagaaatagg caaagcaacc
     6841 gcaaaatatt tcttttactc taatatcatg aacttcttca aaacagaaat tacacttgca
     6901 aatggagaga ttcgcaaacg ccctctaatc gaaactaatg gggaaactgg agaaattgtc
     6961 tgggataaag ggcgagattt tgccacagtg cgcaaagtat tgtccatgcc ccaagtcaat
     7021 attgtcaaga aaacagaagt acagacaggc ggattctcca aggagtcaat tttaccaaaa
     7081 agaaattcgg acaagcttat tgctcgtaaa aaagactggg atccaaaaaa atatggtggt
     7141 tttgatagtc caacggtagc ttattcagtc ctagtggttg ctaaggtgga aaaagggaaa
     7201 tcgaagaagt taaaatccgt taaagagtta ctagggatca caattatgga aagaagttcc
     7261 tttgaaaaaa atccgattga ctttttagaa gctaaaggat ataaggaagt taaaaaagac
     7321 ttaatcatta aactacctaa atatagtctt tttgagttag aaaacggtcg taaacggatg
     7381 ctggctagtg ccggagaatt acaaaaagga aatgagctgg ctctgccaag caaatatgtg
     7441 aattttttat atttagctag tcattatgaa aagttgaagg gtagtccaga agataacgaa
     7501 caaaaacaat tgtttgtgga gcagcataag cattatttag atgagattat tgagcaaatc
     7561 agtgaatttt ctaagcgtgt tattttagca gatgccaatt tagataaagt tcttagtgca
     7621 tataacaaac atagagacaa accaatacgt gaacaagcag aaaatattat tcatttattt
     7681 acgttgacga atcttggagc tcccgctgct tttaaatatt ttgatacaac aattgatcgt
     7741 aaacgatata cgtctacaaa agaagtttta gatgccactc ttatccatca atccatcact
     7801 ggtctttatg aaacacgcat tgatttgagt cagctaggag gtgacgggtc acctaagaaa
     7861 aaacgaaaag ttgaggatcc taaaaagaaa cgaaaagttg ataccggtta ataacattgg
     7921 aagtggataa cggatccgcg atcgcggcgc gccacctggt ggccggccgg taccacgcgt
     7981 gcgcgctgat ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc
     8041 tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct
     8101 gaaaggagga actatatccg gatatccaca ggacgggtgt ggtcgccatg atcgcgtagt
     8161 cgatagtggc tccaagtagc gaagcgagca ggactgggcg gcggccaaag cggtcggaca
     8221 gtgctccgag aacgggtgcg catagaaatt gcatcaacgc atatagcgct agcagcacgc
     8281 catagtgact ggcgatgctg tcggaatgga cgatatcccg caagaggccc ggcagtaccg
     8341 gcataaccaa gcctatgcct acagcatcca gggtgacggt gccgaggatg acgatgagcg
     8401 cattgttaga tttcatacac ggtgcctgac tgcgttagca atttaactgt gataaactac
     8461 cgcattaaag cttatcgatg ataagctgtc aaacatgaga attcttgaag acgaaagggc
     8521 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     8581 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     8641 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata acattgaaaa
     8701 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     8761 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     8821 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     8881 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     8941 gtattatccc gtgttgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     9001 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     9061 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     9121 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     9181 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     9241 accacgatgc ctgcagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     9301 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     9361 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     9421 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     9481 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     9541 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     9601 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     9661 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     9721 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     9781 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     9841 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     9901 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     9961 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
    10021 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
    10081 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gct
//
