LOCUS       sequence-91657-e       13793 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cas9 driven by double 35S, BAR for plant selection, I-PpoI site to 
            accept gRNA from pUC gRNA Shuttle.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13793)
  AUTHORS   Jacobs TB, LaFayette PR, Schmitz RJ, Parrott WA
  TITLE     Targeted genome modifications in soybean with CRISPR/Cas9.
  JOURNAL   BMC Biotechnol. 2015 Mar 12;15:16. doi: 10.1186/s12896-015-0131-2.
  PUBMED    25879861
REFERENCE   2  (bases 1 to 13793)
  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: "BMC 
            Biotechnol."; date: "2015-03-12"; volume: "15:16. doi"; pages: " 
            10.1186/s12896-015-0131-2"
COMMENT     Sequence Label: p201B Cas9
FEATURES             Location/Qualifiers
     source          1..13793
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1187..1816
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             2250..3317
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="GRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESWQA
                     AADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLSKR
                     DRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKPGR
                     VFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGEAL
                     ISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYRLA
                     RRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPILV
                     MRYRNLIEGEASAGS"
     rep_origin      3383..3577
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(3813..3835)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3921..4061
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3976..3995
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4076..4093)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4247..4835)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4327..4346)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4930..5745)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(4930..5745)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     5653..5672
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5737..5756
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     misc_feature    6038..6062
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     protein_bind    6239..6260
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6275..6280,6281..6298,6299..6305)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6313..6329
                     /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)."
     primer_bind     6318..6340
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6337..6353
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6337..6353
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     terminator      complement(6409..6661)
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     CDS             complement(6693..7244)
                     /codon_start=1
                     /gene="Streptomyces hygroscopicus bar"
                     /product="phosphinothricin acetyltransferase"
                     /label=BlpR
                     /note="confers resistance to bialophos or phosphinothricin"
                     /translation="MSPERRPADIRRATEADMPAVCTIVNHYIETSTVNFRTEPQEPQE
                     WTDDLVRLRERYPWLVAEVDGEVAGIAYAGPWKARNAYDWTAESTVYVSPRHQRTGLGS
                     TLYTHLLKSLEAQGFKSVVAVIGLPNDPSVRMHEALGYAPRGMLRAAGFKHGNWHDVGF
                     WQLDFSLPVPPRPVLPVTEI"
     promoter        complement(7295..7971)
                     /label=CaMV 35S promoter (enhanced)
                     /note="cauliflower mosaic virus 35S promoter with a 
                     duplicated enhancer region"
     promoter        8225..8901
                     /label=CaMV 35S promoter (enhanced)
                     /note="cauliflower mosaic virus 35S promoter with a 
                     duplicated enhancer region"
     CDS             8947..13050
                     /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             8947..13050
                     /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             8947..13050
                     /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             13063..13083
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      13150..13402
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     primer_bind     complement(13444..13461)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(13444..13460)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(13453..13475)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     misc_feature    13656..13680
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
ORIGIN
        1 agtactttga tccaacccct ccgctgctat agtgcagtcg gcttctgacg ttcagtgcag
       61 ccgtcttctg aaaacgacat gtcgcacaag tcctaagtta cgcgacaggc tgccgccctg
      121 cccttttcct ggcgttttct tgtcgcgtgt tttagtcgca taaagtagaa tacttgcgac
      181 tagaaccgga gacattacgc catgaacaag agcgccgccg ctggcctgct gggctatgcc
      241 cgcgtcagca ccgacgacca ggacttgacc aaccaacggg ccgaactgca cgcggccggc
      301 tgcaccaagc tgttttccga gaagatcacc ggcaccaggc gcgaccgccc ggagctggcc
      361 aggatgcttg accacctacg ccctggcgac gttgtgacag tgaccaggct agaccgcctg
      421 gcccgcagca cccgcgacct actggacatt gccgagcgca tccaggaggc cggcgcgggc
      481 ctgcgtagcc tggcagagcc gtgggccgac accaccacgc cggccggccg catggtgttg
      541 accgtgttcg ccggcattgc cgagttcgag cgttccctaa tcatcgaccg cacccggagc
      601 gggcgcgagg ccgccaaggc ccgaggcgtg aagtttggcc cccgccctac cctcaccccg
      661 gcacagatcg cgcacgcccg cgagctgatc gaccaggaag gccgcaccgt gaaagaggcg
      721 gctgcactgc ttggcgtgca tcgctcgacc ctgtaccgcg cacttgagcg cagcgaggaa
      781 gtgacgccca ccgaggccag gcggcgcggt gccttccgtg aggacgcatt gaccgaggcc
      841 gacgccctgg cggccgccga gaatgaacgc caagaggaac aagcatgaaa ccgcaccagg
      901 acggccagga cgaaccgttt ttcattaccg aagagatcga ggcggagatg atcgcggccg
      961 ggtacgtgtt cgagccgccc gcgcacgtct caaccgtgcg gctgcatgaa atcctggccg
     1021 gtttgtctga tgccaagctg gcggcctggc cggccagctt ggccgctgaa gaaaccgagc
     1081 gccgccgtct aaaaaggtga tgtgtatttg agtaaaacag cttgcgtcat gcggtcgctg
     1141 cgtatatgat gcgatgagta aataaacaaa tacgcaaggg gaacgcatga aggttatcgc
     1201 tgtacttaac cagaaaggcg ggtcaggcaa gacgaccatc gcaacccatc tagcccgcgc
     1261 cctgcaactc gccggggccg atgttctgtt agtcgattcc gatccccagg gcagtgcccg
     1321 cgattgggcg gccgtgcggg aagatcaacc gctaaccgtt gtcggcatcg accgcccgac
     1381 gattgaccgc gacgtgaagg ccatcggccg gcgcgacttc gtagtgatcg acggagcgcc
     1441 ccaggcggcg gacttggctg tgtccgcgat caaggcagcc gacttcgtgc tgattccggt
     1501 gcagccaagc ccttacgaca tatgggccac cgccgacctg gtggagctgg ttaagcagcg
     1561 cattgaggtc acggatggaa ggctacaagc ggcctttgtc gtgtcgcggg cgatcaaagg
     1621 cacgcgcatc ggcggtgagg ttgccgaggc gctggccggg tacgagctgc ccattcttga
     1681 gtcccgtatc acgcagcgcg tgagctaccc aggcactgcc gccgccggca caaccgttct
     1741 tgaatcagaa cccgagggcg acgctgcccg cgaggtccag gcgctggccg ctgaaattaa
     1801 atcaaaactc atttgagtta atgaggtaaa gagaaaatga gcaaaagcac aaacacgcta
     1861 agtgccggcc gtccgagcgc acgcagcagc aaggctgcaa cgttggccag cctggcagac
     1921 acgccagcca tgaagcgggt caactttcag ttgccggcgg aggatcacac caagctgaag
     1981 atgtacgcgg tacgccaagg caagaccatt accgagctgc tatctgaata catcgcgcag
     2041 ctaccagagt aaatgagcaa atgaataaat gagtagatga attttagcgg ctaaaggagg
     2101 cggcatggaa aatcaagaac aaccaggcac cgacgccgtg gaatgcccca tgtgtggagg
     2161 aacgggcggt tggccaggcg taagcggctg ggttgtctgc cggccctgca atggcactgg
     2221 aacccccaag cccgaggaat cggcgtgacg gtcgcaaacc atccggcccg gtacaaatcg
     2281 gcgcggcgct gggtgatgac ctggtggaga agttgaaggc cgcgcaggcc gcccagcggc
     2341 aacgcatcga ggcagaagca cgccccggtg aatcgtggca agcggccgct gatcgaatcc
     2401 gcaaagaatc ccggcaaccg ccggcagccg gtgcgccgtc gattaggaag ccgcccaagg
     2461 gcgacgagca accagatttt ttcgttccga tgctctatga cgtgggcacc cgcgatagtc
     2521 gcagcatcat ggacgtggcc gttttccgtc tgtcgaagcg tgaccgacga gctggcgagg
     2581 tgatccgcta cgagcttcca gacgggcacg tagaggtttc cgcagggccg gccggcatgg
     2641 ccagtgtgtg ggattacgac ctggtactga tggcggtttc ccatctaacc gaatccatga
     2701 accgataccg ggaagggaag ggagacaagc ccggccgcgt gttccgtcca cacgttgcgg
     2761 acgtactcaa gttctgccgg cgagccgatg gcggaaagca gaaagacgac ctggtagaaa
     2821 cctgcattcg gttaaacacc acgcacgttg ccatgcagcg tacgaagaag gccaagaacg
     2881 gccgcctggt gacggtatcc gagggtgaag ccttgattag ccgctacaag atcgtaaaga
     2941 gcgaaaccgg gcggccggag tacatcgaga tcgagctagc tgattggatg taccgcgaga
     3001 tcacagaagg caagaacccg gacgtgctga cggttcaccc cgattacttt ttgatcgatc
     3061 ccggcatcgg ccgttttctc taccgcctgg cacgccgcgc cgcaggcaag gcagaagcca
     3121 gatggttgtt caagacgatc tacgaacgca gtggcagcgc cggagagttc aagaagttct
     3181 gtttcaccgt gcgcaagctg atcgggtcaa atgacctgcc ggagtacgat ttgaaggagg
     3241 aggcggggca ggctggcccg atcctagtca tgcgctaccg caacctgatc gagggcgaag
     3301 catccgccgg ttcctaatgt acggagcaga tgctagggca aattgcccta gcaggggaaa
     3361 aaggtcgaaa aggtctcttt cctgtggata gcacgtacat tgggaaccca aagccgtaca
     3421 ttgggaaccg gaacccgtac attgggaacc caaagccgta cattgggaac cggtcacaca
     3481 tgtaagtgac tgatataaaa gagaaaaaag gcgatttttc cgcctaaaac tctttaaaac
     3541 ttattaaaac tcttaaaacc cgcctggcct gtgcataact gtctggccag cgcacagccg
     3601 aagagctgca aaaagcgcct acccttcggt cgctgcgctc cctacgcccc gccgcttcgc
     3661 gtcggcctat cgcggccgct ggccgctcaa aaatggctgg cctacggcca ggcaatctac
     3721 cagggcgcgg acaagccgcg ccgtcgccac tcgaccgccg gcgcccacat caaggcaccc
     3781 tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
     3841 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
     3901 ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag cggagtgtat
     3961 actggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg
     4021 aaataccgca cagatgcgta aggagaaaat accgcatcag gcgctcttcc gcttcctcgc
     4081 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     4141 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     4201 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     4261 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     4321 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     4381 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     4441 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     4501 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     4561 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4621 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4681 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4741 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4801 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     4861 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg caggatcatg
     4921 aattaattct tagaaaaact catcgagcat caaatgaaac tgcaatttat tcatatcagg
     4981 attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa actcaccgag
     5041 gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc gtccaacatc
     5101 aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg
     5161 agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc agacttgttc
     5221 aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac cgttattcat
     5281 tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac aattacaaac
     5341 aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat tttcacctga
     5401 atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag tggtgagtaa
     5461 ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca taaattccgt
     5521 cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac ctttgccatg
     5581 tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg tcgcacctga
     5641 ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca tgttggaatt
     5701 taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcataacac cccttgtatt
     5761 actgtttatg taagcagaca gttttattgt tcatgatctg gatcacaggc agcaacgctc
     5821 tgtcatcgtt acaatcaaca tgctaccctc cgcgagatca tccgtgtttc aaacccggca
     5881 gcttagttgc cgttcttccg aatagcatcg gtaacatgag caaagtctgc cgccttacaa
     5941 cggctctccc gctgacgccg tcccggactg atgggctgcc tgtatcgagt ggtgattttg
     6001 tgccgagctg ccggtcgggg agctgttggc tggctggtgg caggatatat tgtggtgtaa
     6061 acaaattgac gcttagacaa cttaataaca cattgcggac gtttttaatg tactgaatta
     6121 acgccgaatt gctctagcca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     6181 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     6241 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     6301 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gacatgatta
     6361 cgaattcctt gactagaggg taggcgcggc ttaattaaat caattcccga tctagtaaca
     6421 tagatgacac cgcgcgcgat aatttatcct agtttgcgcg ctatattttg ttttctatcg
     6481 cgtattaaat gtataattgc gggactctaa tcataaaaac ccatctcata aataacgtca
     6541 tgcattacat gttaattatt acatgcttaa cgtaattcaa cagaaattat atgataatca
     6601 tcgcaagacc ggcaacagga ttcaatctta agaaacttta ttgccaaatg tttgaacgat
     6661 cggggaaatt cgagctcggc cggcctcgag atttaaatct cggtgacggg caggaccgga
     6721 cggggcggta ccggcaggct gaagtccagc tgccagaaac ccacgtcatg ccagttcccg
     6781 tgcttgaagc cggccgcccg cagcatgccg cggggggcat atccgagcgc ctcgtgcatg
     6841 cgcacgctcg ggtcgttggg cagcccgatg acagcgacca cgctcttgaa gccctgtgcc
     6901 tccagggact tcagcaggtg ggtgtagagc gtggagccca gtcccgtccg ctggtggcgg
     6961 ggggagacgt acacggtcga ctcggccgtc cagtcgtagg cgttgcgtgc cttccagggg
     7021 cccgcgtagg cgatgccggc gacctcgccg tccacctcgg cgacgagcca gggatagcgc
     7081 tcccgcagac ggacgaggtc gtccgtccac tcctgcggtt cctgcggctc ggtacggaag
     7141 ttgaccgtgc ttgtctcgat gtagtggttg acgatggtgc agaccgccgg catgtccgcc
     7201 tcggtggcac ggcggatgtc ggccgggcgt cgttctgggc tcatcctagg ctagcggtcg
     7261 agagagatag atttgtagag agagactggt gatttcagcg tgtcctctcc aaatgaaatg
     7321 aacttcctta tatagaggaa gggtcttgcg aaggatagtg ggattgtgcg tcatccctta
     7381 cgtcagtgga gatatcacat caatccactt gctttgaaga cgtggttgga acgtcttctt
     7441 tttccacgat gctcctcgtg ggtgggggtc catctttggg accactgtcg gcagaggcat
     7501 cttgaacgat agcctttcct ttatcgcaat gatggcattt gtaggtgcca ccttcctttt
     7561 ctactgtcct tttgatgaag tgacagatag ctgggcaatg gaatccgagg aggtttcccg
     7621 atattaccct ttgttgaaaa gtctcaatag ccctttggtc ttctgagact gtatctttga
     7681 tattcttgga gtagacgaga gtgtcgtgct ccaccatgtt cacatcaatc cacttgcttt
     7741 gaagacgtgg ttggaacgtc ttctttttcc acgatgctcc tcgtgggtgg gggtccatct
     7801 ttgggaccac tgtcggcaga ggcatcttga acgatagcct ttcctttatc gcaatgatgg
     7861 catttgtagg tgccaccttc cttttctact gtccttttga tgaagtgaca gatagctggg
     7921 caatggaatc cgaggaggtt tcccgatatt accctttgtt gaaaagtctc aatagccctt
     7981 tggtcttctg agactgtatc tttgatattc ttggagtaga cgagagtgtc gtgctccacc
     8041 atgttgggac tagtgagtcg tattacaatt catgactctc ttaaggtagc caaaatcgat
     8101 atctagggat aacagggtaa tcgatcacta gaattcggcg cgccgggccc aacatggtgg
     8161 agcacgacac tctcgtctac tccaagaata tcaaagatac agtctcagaa gaccaaaggg
     8221 ctattgagac ttttcaacaa agggtaatat cgggaaacct cctcggattc cattgcccag
     8281 ctatctgtca cttcatcaaa aggacagtag aaaaggaagg tggcacctac aaatgccatc
     8341 attgcgataa aggaaaggct atcgttcaag atgcctctgc cgacagtggt cccaaagatg
     8401 gacccccacc cacgaggagc atcgtggaaa aagaagacgt tccaaccacg tcttcaaagc
     8461 aagtggattg atgtgaacat ggtggagcac gacactctcg tctactccaa gaatatcaaa
     8521 gatacagtct cagaagacca aagggctatt gagacttttc aacaaagggt aatatcggga
     8581 aacctcctcg gattccattg cccagctatc tgtcacttca tcaaaaggac agtagaaaag
     8641 gaaggtggca cctacaaatg ccatcattgc gataaaggaa aggctatcgt tcaagatgcc
     8701 tctgccgaca gtggtcccaa agatggaccc ccacccacga ggagcatcgt ggaaaaagaa
     8761 gacgttccaa ccacgtcttc aaagcaagtg gattgatgtg atatctccac tgacgtaagg
     8821 gatgacgcac aatcccacta tccttcgcaa gacccttcct ctatataagg aagttcattt
     8881 catttggaga ggacacgctg aaatcaccag tctctctcta caaatctatc tctctcgacc
     8941 gctagcatgg acaagaagta ctccattggg ctcgatatcg gcacaaacag cgtcggctgg
     9001 gccgtcatta cggacgagta caaggtgccg agcaaaaaat tcaaagttct gggcaatacc
     9061 gatcgccaca gcataaagaa gaacctcatt ggcgccctcc tgttcgactc cggggagacg
     9121 gccgaagcca cgcggctcaa aagaacagca cggcgcagat atacccgcag aaagaatcgg
     9181 atctgctacc tgcaggagat ctttagtaat gagatggcta aggtggatga ctctttcttc
     9241 cataggctgg aggagtcctt tttggtggag gaggataaaa agcacgagcg ccacccaatc
     9301 tttggcaata tcgtggacga ggtggcgtac catgaaaagt acccaaccat atatcatctg
     9361 aggaagaagc ttgtagacag tactgataag gctgacttgc ggttgatcta tctcgcgctg
     9421 gcgcatatga tcaaatttcg gggacacttc ctcatcgagg gggacctgaa cccagacaac
     9481 agcgatgtcg acaaactctt tatccaactg gttcagactt acaatcagct tttcgaagag
     9541 aacccgatca acgcatccgg agttgacgcc aaagcaatcc tgagcgctag gctgtccaaa
     9601 tcccggcggc tcgaaaacct catcgcacag ctccctgggg agaagaagaa cggcctgttt
     9661 ggtaatctta tcgccctgtc actcgggctg acccccaact ttaaatctaa cttcgacctg
     9721 gccgaagatg ccaagcttca actgagcaaa gacacctacg atgatgatct cgacaatctg
     9781 ctggcccaga tcggcgacca gtacgcagac ctttttttgg cggcaaagaa cctgtcagac
     9841 gccattctgc tgagtgatat tctgcgagtg aacacggaga tcaccaaagc tccgctgagc
     9901 gctagtatga tcaagcgcta tgatgagcac caccaagact tgactttgct gaaggccctt
     9961 gtcagacagc aactgcctga gaagtacaag gaaattttct tcgatcagtc taaaaatggc
    10021 tacgccggat acattgacgg cggagcaagc caggaggaat tttacaaatt tattaagccc
    10081 atcttggaaa aaatggacgg caccgaggag ctgctggtaa agcttaacag agaagatctg
    10141 ttgcgcaaac agcgcacttt cgacaatgga agcatccccc accagattca cctgggcgaa
    10201 ctgcacgcta tcctcaggcg gcaagaggat ttctacccct ttttgaaaga taacagggaa
    10261 aagattgaga aaatcctcac atttcggata ccctactatg taggccccct cgcccgggga
    10321 aattccagat tcgcgtggat gactcgcaaa tcagaagaga ccatcactcc ctggaacttc
    10381 gaggaagtcg tggataaggg ggcctctgcc cagtccttca tcgaaaggat gactaacttt
    10441 gataaaaatc tgcctaacga aaaggtgctt cctaaacact ctctgctgta cgagtacttc
    10501 acagtttata acgagctcac caaggtcaaa tacgtcacag aagggatgag aaagccagca
    10561 ttcctgtctg gagagcagaa gaaagctatc gtggacctcc tcttcaagac gaaccggaaa
    10621 gttaccgtga aacagctcaa agaagactat ttcaaaaaga ttgaatgttt cgactctgtt
    10681 gaaatcagcg gagtggagga tcgcttcaac gcatccctgg gaacgtatca cgatctcctg
    10741 aaaatcatta aagacaagga cttcctggac aatgaggaga acgaggacat tcttgaggac
    10801 attgtcctca cccttacgtt gtttgaagat agggagatga ttgaagaacg cttgaaaact
    10861 tacgctcatc tcttcgacga caaagtcatg aaacagctca agaggcgccg atatacagga
    10921 tgggggcggc tgtcaagaaa actgatcaat gggatccgag acaagcagag tggaaagaca
    10981 atcctggatt ttcttaagtc cgatggattt gccaaccgga acttcatgca gttgatccat
    11041 gatgactctc tcacctttaa ggaggacatc cagaaagcac aagtttctgg ccagggggac
    11101 agtcttcacg agcacatcgc taatcttgca ggtagcccag ctatcaaaaa gggaatactg
    11161 cagaccgtta aggtcgtgga tgaactcgtc aaagtaatgg gaaggcataa gcccgagaat
    11221 atcgttatcg agatggcccg agagaaccaa actacccaga agggacagaa gaacagtagg
    11281 gaaaggatga agaggattga agagggtata aaagaactgg ggtcccaaat ccttaaggaa
    11341 cacccagttg aaaacaccca gcttcagaat gagaagctct acctgtacta cctgcagaac
    11401 ggcagggaca tgtacgtgga tcaggaactg gacatcaatc ggctctccga ctacgacgtg
    11461 gatcatatcg tgccccagtc ttttctcaaa gatgattcta ttgataataa agtgttgaca
    11521 agatccgata aaaatagagg gaagagtgat aacgtcccct cagaagaagt tgtcaagaaa
    11581 atgaaaaatt attggcggca gctgctgaac gccaaactga tcacacaacg gaagttcgat
    11641 aatctgacta aggctgaacg aggtggcctg tctgagttgg ataaagccgg cttcatcaaa
    11701 aggcagcttg ttgagacacg ccagatcacc aagcacgtgg cccaaattct cgattcacgc
    11761 atgaacacca agtacgatga aaatgacaaa ctgattcgag aggtgaaagt tattactctg
    11821 aagtctaagc tggtctcaga tttcagaaag gactttcagt tttataaggt gagagagatc
    11881 aacaattacc accatgcgca tgatgcctac ctgaatgcag tggtaggcac tgcacttatc
    11941 aaaaaatatc ccaagcttga atctgaattt gtttacggag actataaagt gtacgatgtt
    12001 aggaaaatga tcgcaaagtc tgagcaggaa ataggcaagg ccaccgctaa gtacttcttt
    12061 tacagcaata ttatgaattt tttcaagacc gagattacac tggccaatgg agagattcgg
    12121 aagcgaccac ttatcgaaac aaacggagaa acaggagaaa tcgtgtggga caagggtagg
    12181 gatttcgcga cagtccggaa ggtcctgtcc atgccgcagg tgaacatcgt taaaaagacc
    12241 gaagtacaga ccggaggctt ctccaaggaa agtatcctcc cgaaaaggaa cagcgacaag
    12301 ctgatcgcac gcaaaaaaga ttgggacccc aagaaatacg gcggattcga ttctcctaca
    12361 gtcgcttaca gtgtactggt tgtggccaaa gtggagaaag ggaagtctaa aaaactcaaa
    12421 agcgtcaagg aactgctggg catcacaatc atggagcgat caagcttcga aaaaaacccc
    12481 atcgactttc tcgaggcgaa aggatataaa gaggtcaaaa aagacctcat cattaagctt
    12541 cccaagtact ctctctttga gcttgaaaac ggccggaaac gaatgctcgc tagtgcgggc
    12601 gagctgcaga aaggtaacga gctggcactg ccctctaaat acgttaattt cttgtatctg
    12661 gccagccact atgaaaagct caaagggtct cccgaagata atgagcagaa gcagctgttc
    12721 gtggaacaac acaaacacta ccttgatgag atcatcgagc aaataagcga attctccaaa
    12781 agagtgatcc tcgccgacgc taacctcgat aaggtgcttt ctgcttacaa taagcacagg
    12841 gataagccca tcagggagca ggcagaaaac attatccact tgtttactct gaccaacttg
    12901 ggcgcgcctg cagccttcaa gtacttcgac accaccatag acagaaagcg gtacacctct
    12961 acaaaggagg tcctggacgc cacactgatt catcagtcaa ttacggggct ctatgaaaca
    13021 agaatcgacc tctctcagct cggtggagac agcagggctg accccaagaa gaagaggaag
    13081 gtgtgaccgc ggatatctgc agaagcttac gcgtcgacgt ctcgaggccg gccgagctcg
    13141 aatttccccg atcgttcaaa catttggcaa taaagtttct taagattgaa tcctgttgcc
    13201 ggtcttgcga tgattatcat ataatttctg ttgaattacg ttaagcatgt aataattaac
    13261 atgtaatgca tgacgttatt tatgagatgg gtttttatga ttagagtccc gcaattatac
    13321 atttaatacg cgatagaaaa caaaatatag cgcgcaaact aggataaatt atcgcgcgcg
    13381 gtgtcatcta tgttactaga tcgggaattg atatcgtagg gataacaggg taataagctt
    13441 ggcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
    13501 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
    13561 tcgcccttcc caacagttgc gcagcctgaa tggcgaatga gcttgagctt ggatcagatt
    13621 gtcgtttccc gccttcagtt taaactatca gtgtttgaca ggatatattg gcgggtaaac
    13681 ctaagagaaa agagcgttta ttagaataac ggatatttaa aagggcgtga aaaggtttat
    13741 ccgttcgtcc atttgtatgt gcatgccaac cacagggttc ccctcgggat caa
//
