LOCUS       sequence-329607-       11755 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for constituive spCas9 and tet-inducible SEO1 targeting 
            sgRNA expression for double stranded break formation in yeast.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11755)
  TITLE     Burgess Lab Plasmids
REFERENCE   2  (bases 1 to 11755)
  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"
COMMENT     Sequence Label: pNSU299_SEO1
FEATURES             Location/Qualifiers
     source          1..11755
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..399
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     primer_bind     403..419
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     403..419
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             431..4534
                     /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             431..4534
                     /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             431..4534
                     /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             4547..4567
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      4583..4830
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4599..4617)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(4848..4867)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4849..4867)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4877..4894)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4877..4893)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4886..4908)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5034..5489
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5121..5140)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5331..5352
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5592..6266)
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     promoter        complement(6267..6548)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(6569..6588)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6688..6710
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6748..6766)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6834..6938
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6939..7007,7008..7799)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(7157..7176)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7970..8558
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8459..8478
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    8942..8960
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_RNA        9046..9121
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      9134..9196
                     /label=RPR1 terminator
                     /note="transcription terminator for the S. cerevisiae RNase
                     P RNA gene"
     CDS             10331..10954
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(10404..10423)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     11451..11468
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    11585..11606
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(11621..11626,11627..11644,11645..11651)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11659..11675
                     /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     11664..11686
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     11683..11699
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     11683..11699
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     11718..11738
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        11720..11738
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 catagcttca aaatgtttct actccttttt tactcttcca gattttctcg gactccgcgc
       61 atcgccgtac cacttcaaaa cacccaagca cagcatacta aatttcccct ctttcttcct
      121 ctagggtgtc gttaattacc cgtactaaag gtttggaaaa gaaaaaagag accgcctcgt
      181 ttctttttct tcgtcgaaaa aggcaataaa aatttttatc acgtttcttt ttcttgaaaa
      241 tttttttttt gatttttttc tctttcgatg acctcccatt gatatttaag ttaataaacg
      301 gtcttcaatt tctcaagttt cagtttcatt tttcttgttc tattacaact ttttttactt
      361 cttgctcatt agaaagaaag catagcaatc taatctaagt tttctagaac tagtggatcc
      421 cccgggaaaa atggacaaga agtactccat tgggctcgat atcggcacaa acagcgtcgg
      481 ttgggccgtc attacggacg agtacaaggt gccgagcaaa aaattcaaag ttctgggcaa
      541 taccgatcgc cacagcataa agaagaacct cattggcgcc ctcctgttcg actccgggga
      601 gacggccgaa gccacgcggc tcaaaagaac agcacggcgc agatataccc gcagaaagaa
      661 tcggatctgc tacctgcagg agatctttag taatgagatg gctaaggtgg atgactcttt
      721 cttccatagg ctggaggagt cctttttggt ggaggaggat aaaaagcacg agcgccaccc
      781 aatctttggc aatatcgtgg acgaggtggc gtaccatgaa aagtacccaa ccatatatca
      841 tctgaggaag aagcttgtag acagtactga taaggctgac ttgcggttga tctatctcgc
      901 gctggcgcat atgatcaaat ttcggggaca cttcctcatc gagggggacc tgaacccaga
      961 caacagcgat gtcgacaaac tctttatcca actggttcag acttacaatc agcttttcga
     1021 agagaacccg atcaacgcat ccggagttga cgccaaagca atcctgagcg ctaggctgtc
     1081 caaatcccgg cggctcgaaa acctcatcgc acagctccct ggggagaaga agaacggcct
     1141 gtttggtaat cttatcgccc tgtcactcgg gctgaccccc aactttaaat ctaacttcga
     1201 cctggccgaa gatgccaagc ttcaactgag caaagacacc tacgatgatg atctcgacaa
     1261 tctgctggcc cagatcggcg accagtacgc agaccttttt ttggcggcaa agaacctgtc
     1321 agacgccatt ctgctgagtg atattctgcg agtgaacacg gagatcacca aagctccgct
     1381 gagcgctagt atgatcaagc gctatgatga gcaccaccaa gacttgactt tgctgaaggc
     1441 ccttgtcaga cagcaactgc ctgagaagta caaggaaatt ttcttcgatc agtctaaaaa
     1501 tggctacgcc ggatacattg acggcggagc aagccaggag gaattttaca aatttattaa
     1561 gcccatcttg gaaaaaatgg acggcaccga ggagctgctg gtaaagctta acagagaaga
     1621 tctgttgcgc aaacagcgca ctttcgacaa tggaagcatc ccccaccaga ttcacctggg
     1681 cgaactgcac gctatcctca ggcggcaaga ggatttctac ccctttttga aagataacag
     1741 ggaaaagatt gagaaaatcc tcacatttcg gataccctac tatgtaggcc ccctcgcccg
     1801 gggaaattcc agattcgcgt ggatgactcg caaatcagaa gagaccatca ctccctggaa
     1861 cttcgaggaa gtcgtggata agggggcctc tgcccagtcc ttcatcgaaa ggatgactaa
     1921 ctttgataaa aatctgccta acgaaaaggt gcttcctaaa cactctctgc tgtacgagta
     1981 cttcacagtt tataacgagc tcaccaaggt caaatacgtc acagaaggga tgagaaagcc
     2041 agcattcctg tctggagagc agaagaaagc tatcgtggac ctcctcttca agacgaaccg
     2101 gaaagttacc gtgaaacagc tcaaagaaga ctatttcaaa aagattgaat gtttcgactc
     2161 tgttgaaatc agcggagtgg aggatcgctt caacgcatcc ctgggaacgt atcacgatct
     2221 cctgaaaatc attaaagaca aggacttcct ggacaatgag gagaacgagg acattcttga
     2281 ggacattgtc ctcaccctta cgttgtttga agatagggag atgattgaag aacgcttgaa
     2341 aacttacgct catctcttcg acgacaaagt catgaaacag ctcaagaggc gccgatatac
     2401 aggatggggg cggctgtcaa gaaaactgat caatgggatc cgagacaagc agagtggaaa
     2461 gacaatcctg gattttctta agtccgatgg atttgccaac cggaacttca tgcagttgat
     2521 ccatgatgac tctctcacct ttaaggagga catccagaaa gcacaagttt ctggccaggg
     2581 ggacagtctt cacgagcaca tcgctaatct tgcaggtagc ccagctatca aaaagggaat
     2641 actgcagacc gttaaggtcg tggatgaact cgtcaaagta atgggaaggc ataagcccga
     2701 gaatatcgtt atcgagatgg cccgagagaa ccaaactacc cagaagggac agaagaacag
     2761 tagggaaagg atgaagagga ttgaagaggg tataaaagaa ctggggtccc aaatccttaa
     2821 ggaacaccca gttgaaaaca cccagcttca gaatgagaag ctctacctgt actacctgca
     2881 gaacggcagg gacatgtacg tggatcagga actggacatc aatcggctct ccgactacga
     2941 cgtggatcat atcgtgcccc agtcttttct caaagatgat tctattgata ataaagtgtt
     3001 gacaagatcc gataaaaata gagggaagag tgataacgtc ccctcagaag aagttgtcaa
     3061 gaaaatgaaa aattattggc ggcagctgct gaacgccaaa ctgatcacac aacggaagtt
     3121 cgataatctg actaaggctg aacgaggtgg cctgtctgag ttggataaag ccggcttcat
     3181 caaaaggcag cttgttgaga cacgccagat caccaagcac gtggcccaaa ttctcgattc
     3241 acgcatgaac accaagtacg atgaaaatga caaactgatt cgagaggtga aagttattac
     3301 tctgaagtct aagctggtct cagatttcag aaaggacttt cagttttata aggtgagaga
     3361 gatcaacaat taccaccatg cgcatgatgc ctacctgaat gcagtggtag gcactgcact
     3421 tatcaaaaaa tatcccaagc ttgaatctga atttgtttac ggagactata aagtgtacga
     3481 tgttaggaaa atgatcgcaa agtctgagca ggaaataggc aaggccaccg ctaagtactt
     3541 cttttacagc aatattatga attttttcaa gaccgagatt acactggcca atggagagat
     3601 tcggaagcga ccacttatcg aaacaaacgg agaaacagga gaaatcgtgt gggacaaggg
     3661 tagggatttc gcgacagtcc ggaaggtcct gtccatgccg caggtgaaca tcgttaaaaa
     3721 gaccgaagta cagaccggag gcttctccaa ggaaagtatc ctcccgaaaa ggaacagcga
     3781 caagctgatc gcacgcaaaa aagattggga ccccaagaaa tacggcggat tcgattctcc
     3841 tacagtcgct tacagtgtac tggttgtggc caaagtggag aaagggaagt ctaaaaaact
     3901 caaaagcgtc aaggaactgc tgggcatcac aatcatggag cgatcaagct tcgaaaaaaa
     3961 ccccatcgac tttctcgagg cgaaaggata taaagaggtc aaaaaagacc tcatcattaa
     4021 gcttcccaag tactctctct ttgagcttga aaacggccgg aaacgaatgc tcgctagtgc
     4081 gggcgagctg cagaaaggta acgagctggc actgccctct aaatacgtta atttcttgta
     4141 tctggccagc cactatgaaa agctcaaagg gtctcccgaa gataatgagc agaagcagct
     4201 gttcgtggaa caacacaaac actaccttga tgagatcatc gagcaaataa gcgaattctc
     4261 caaaagagtg atcctcgccg acgctaacct cgataaggtg ctttctgctt acaataagca
     4321 cagggataag cccatcaggg agcaggcaga aaacattatc cacttgttta ctctgaccaa
     4381 cttgggcgcg cctgcagcct tcaagtactt cgacaccacc atagacagaa agcggtacac
     4441 ctctacaaag gaggtcctgg acgccacact gattcatcag tcaattacgg ggctctatga
     4501 aacaagaatc gacctctctc agctcggtgg agacagcagg gctgacccca agaagaagag
     4561 gaaggtgtga tctcttctcg agtcatgtaa ttagttatgt cacgcttaca ttcacgccct
     4621 ccccccacat ccgctctaac cgaaaaggaa ggagttagac aacctgaagt ctaggtccct
     4681 atttattttt ttatagttat gttagtatta agaacgttat ttatatttca aatttttctt
     4741 ttttttctgt acagacgcgt gtacgcatgt aacattatac tgaaaacctt gcttgagaag
     4801 gttttgggac gctcgaaggc tttaatttgc ggccggtacc caattcgccc tatagtgagt
     4861 cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg
     4921 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag
     4981 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggacgcgcc
     5041 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     5101 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     5161 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     5221 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     5281 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     5341 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     5401 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     5461 ttttaacaaa atattaacgc ttacaatttc ctgatgcggt attttctcct tacgcatctg
     5521 tgcggtattt cacaccgcat agatcggcaa gtgcacaaac aatacttaaa taaatactac
     5581 tcagtaataa cctatttctt agcatttttg acgaaatttg ctattttgtt agagtctttt
     5641 acaccatttg tctccacacc tccgcttaca tcaacaccaa taacgccatt taatctaagc
     5701 gcatcaccaa cattttctgg cgtcagtcca ccagctaaca taaaatgtaa gctttcgggg
     5761 ctctcttgcc ttccaaccca gtcagaaatc gagttccaat ccaaaagttc acctgtccca
     5821 cctgcttctg aatcaaacaa gggaataaac gaatgaggtt tctgtgaagc tgcactgagt
     5881 agtatgttgc agtcttttgg aaatacgagt cttttaataa ctggcaaacc gaggaactct
     5941 tggtattctt gccacgactc atctccatgc agttggacga tatcaatgcc gtaatcattg
     6001 accagagcca aaacatcctc cttaggttga ttacgaaaca cgccaaccaa gtatttcgga
     6061 gtgcctgaac tatttttata tgcttttaca agacttgaaa ttttccttgc aataaccggg
     6121 tcaattgttc tctttctatt gggcacacat ataataccca gcaagtcagc atcggaatct
     6181 agagcacatt ctgcggcctc tgtgctctgc aagccgcaaa ctttcaccaa tggaccagaa
     6241 ctacctgtga aattaataac agacatactc caagctgcct ttgtgtgctt aatcacgtat
     6301 actcacgtgc tcaatagtca ccaatgccct ccctcttggc cctctccttt tcttttttcg
     6361 accgaattaa ttcttaatcg gcaaaaaaag aaaagctccg gatcaagatt gtacgtaagg
     6421 tgacaagcta tttttcaata aagaatatct tccactactg ccatctggcg tcataactgc
     6481 aaagtacaca tatattacga tgctgttcta ttaaatgctt cctatattat atatatagta
     6541 atgtcgttga tctatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     6601 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     6661 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     6721 catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     6781 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     6841 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     6901 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     6961 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     7021 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     7081 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     7141 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     7201 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     7261 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     7321 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     7381 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     7441 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     7501 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     7561 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     7621 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     7681 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     7741 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     7801 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     7861 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     7921 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     7981 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     8041 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     8101 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     8161 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     8221 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     8281 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     8341 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     8401 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     8461 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     8521 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     8581 tacggttcct ggccttttgc tggccttttg ctcgggggat ctgccaattg aacataacat
     8641 ggtagttaca tatactagta atatggttcg gcacacatta aaagtataaa aactatctga
     8701 attacgaatt acatatattg gtcataaaaa tcaatcaatc atcgtgtgtt ttatatgtct
     8761 cttatctaag tataagaata tccatagtta atattcactt acgctacctt ttaacctgta
     8821 atcattgtca acaggatatg ttaacgaccc acattgataa acgctagtat ttctttttcc
     8881 tcttcttatt ggccggctgt ctctatactc ccctatagtc tgtttctttt cgtttcgatt
     8941 gtccctatca gtgatagaga tggcgcacat ggtacgctgt ggtgctcgcg gctgggaacg
     9001 aaactctggg agctgcgatt ggcagaacca aaacacttat ccatggtttt agagctagaa
     9061 atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg
     9121 cttttttctc gagccatatc caacttccaa tttaatcttt cttttttaat tttcacttat
     9181 ttgcgataca gaaagaaaaa agcgatagta actattgaat tttgtttgga tttggttaga
     9241 ttagatatgg tttctcttta tatttacatg ctaaaaatgg gctacaccag agatacataa
     9301 ttagatatat atacgccagt acaccttatc ggcccaagcc ttgtcccaag gcagcgtttt
     9361 gttcttggaa acgctgccct acacgttcgc tatgcttcaa gaacttttct gagcacttca
     9421 tgatgcatgt ttgttcctta ttggttagct ttgatgttgt gaagtcattg acacagtctg
     9481 tgaaacatct ttctaccaga ttagagtaca aacgcatgaa atccttcatt tgcttttgtt
     9541 ccactacttt ttggaactct tgttgttctt tcctgcaggt agtcgtgcaa tgtatgactt
     9601 taagatttgt gagcaggaag aaaagggaga atcttctaac gataaaccct tgaaaaactg
     9661 ggtagactac gctatgttga gttgctacgc aggctgcaca attacacgag aatgctcccg
     9721 cctaggattt aaggctaagg gacgtgcaat gcagacgaca gatctaaatg accgtgtcgg
     9781 tgaagtgttc gccaaacttt tcggttaaca catgcagtga tgcacgcgcg atggtgctaa
     9841 gttacatata tatatatatg tatagccata gtgatgtcta agtaaccttt atggtatatt
     9901 tcttaatgtg gaaagatact agcgcgcgca cccacacaca agcttcgtct tttcttgaag
     9961 aaaagaggaa gctcgctaaa tgggattcca ctttccgttc cctgccagct gatggaaaaa
    10021 ggttagtgga acgatgaaga ataaaaagag agatccactg aggtgaaatt tcagctgaca
    10081 gcgagtttca tgatcgtgat gaacaatggt aacgagttgt ggctgttgcc agggagggtg
    10141 gttctcaact tttaatgtat ggccaaatcg ctacttgggt ttgttatata acaaagaaga
    10201 aataatgaac tgattctctt cctccttctt gtcctttctt aattctgttg taattacctt
    10261 cctttgtaat tttttttgta attattcttc ttaataatcc aaacaaacac acatattaca
    10321 atatattaaa atgtctagat tagataaaag taaagtgatt aacagcgcat tagagctgct
    10381 taatgaggtc ggaatcgaag gtttaacaac ccgtaaactc gcccagaagc taggtgtaga
    10441 gcagcctaca ttgtattggc atgtaaaaaa taagcgggct ttgctcgacg ccttagccat
    10501 tgagatgtta gataggcacc atactcactt ttgcccttta gaaggggaaa gctggcaaga
    10561 ttttttacgt aataacgcta aaagttttag atgtgcttta ctaagtcatc gcgatggagc
    10621 aaaagtacat ttaggtacac ggcctacaga aaaacagtat gaaactctcg aaaatcaatt
    10681 agccttttta tgccaacaag gtttttcact agagaatgca ttatatgcac tcagcgctgt
    10741 ggggcatttt actttaggtt gcgtattgga agatcaagag catcaagtcg ctaaagaaga
    10801 aagggaaaca cctactactg atagtatgcc gccattatta cgacaagcta tcgaattatt
    10861 tgatcaccaa ggtgcagagc cagccttctt attcggcctt gaattgatca tatgcggatt
    10921 agaaaaacaa cttaaatgtg aaagtgggtc ttaagtctga agaatgaatg atttgatgat
    10981 ttctttttcc ctccattttt cttactgaat atatcaatga tatagacttg tatagtttat
    11041 tatttcaaat taagtagcta tatatagtca agataacgtt tgtttgacac gattacatta
    11101 ttcgtcgaca tcttttttca gcctgtcgtg gtagcaattt gaggagtatt attaattgaa
    11161 taggttcatt ttgcgctcgc ataaacagtt ttcgtcaggg acagtatgtt ggaatgagtg
    11221 gtaattaatg gtgacatgac atgttatagc aataaccttg atgtttacat cgtagtttaa
    11281 tgtacacccc gcgaattcgt tcaagtagga gtgcaccaat tgcaaaggga aaagctgaat
    11341 gggcagttcg aaccgcggtc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
    11401 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
    11461 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
    11521 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg
    11581 caattaatgt gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg
    11641 ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc
    11701 atgattacgc caagcgcgca attaaccctc actaaaggga acaaaagctg gagct
//
