LOCUS       sequence-329609-       13093 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for constituive spCas9 and tet-inducible MET16 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 13093)
  TITLE     Burgess Lab Plasmids
REFERENCE   2  (bases 1 to 13093)
  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_MET16
FEATURES             Location/Qualifiers
     source          1..13093
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..624
                     /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(74..93)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     1121..1138
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1255..1276
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1291..1296,1297..1314,1315..1321)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1329..1345
                     /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     1334..1356
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1353..1369
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1353..1369
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1388..1408
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1390..1408
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     3166..3182
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3166..3182
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             3194..7297
                     /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             3194..7297
                     /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             3194..7297
                     /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             7310..7330
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      7346..7593
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(7362..7380)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(7611..7630)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(7612..7630)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(7640..7657)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(7640..7656)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7649..7671)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      7797..8252
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7884..7903)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8094..8115
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(8355..9029)
                     /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(9030..9311)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(9332..9351)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     9451..9473
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(9511..9529)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        9597..9701
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9702..9770,9771..10562)
                     /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(9920..9939)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10733..11321
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     11222..11241
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    11705..11723
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_RNA        11809..11884
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      11897..11959
                     /label=RPR1 terminator
                     /note="transcription terminator for the S. cerevisiae RNase
                     P RNA gene"
ORIGIN
        1 atgtctagat tagataaaag taaagtgatt aacagcgcat tagagctgct taatgaggtc
       61 ggaatcgaag gtttaacaac ccgtaaactc gcccagaagc taggtgtaga gcagcctaca
      121 ttgtattggc atgtaaaaaa taagcgggct ttgctcgacg ccttagccat tgagatgtta
      181 gataggcacc atactcactt ttgcccttta gaaggggaaa gctggcaaga ttttttacgt
      241 aataacgcta aaagttttag atgtgcttta ctaagtcatc gcgatggagc aaaagtacat
      301 ttaggtacac ggcctacaga aaaacagtat gaaactctcg aaaatcaatt agccttttta
      361 tgccaacaag gtttttcact agagaatgca ttatatgcac tcagcgctgt ggggcatttt
      421 actttaggtt gcgtattgga agatcaagag catcaagtcg ctaaagaaga aagggaaaca
      481 cctactactg atagtatgcc gccattatta cgacaagcta tcgaattatt tgatcaccaa
      541 ggtgcagagc cagccttctt attcggcctt gaattgatca tatgcggatt agaaaaacaa
      601 cttaaatgtg aaagtgggtc ttaagtctga agaatgaatg atttgatgat ttctttttcc
      661 ctccattttt cttactgaat atatcaatga tatagacttg tatagtttat tatttcaaat
      721 taagtagcta tatatagtca agataacgtt tgtttgacac gattacatta ttcgtcgaca
      781 tcttttttca gcctgtcgtg gtagcaattt gaggagtatt attaattgaa taggttcatt
      841 ttgcgctcgc ataaacagtt ttcgtcaggg acagtatgtt ggaatgagtg gtaattaatg
      901 gtgacatgac atgttatagc aataaccttg atgtttacat cgtagtttaa tgtacacccc
      961 gcgaattcgt tcaagtagga gtgcaccaat tgcaaaggga aaagctgaat gggcagttcg
     1021 aaccgcggtc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     1081 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     1141 agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
     1201 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt
     1261 gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt
     1321 gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc atgattacgc
     1381 caagcgcgca attaaccctc actaaaggga acaaaagctg gagctcatag cttcaaaatg
     1441 tttctactcc ttttttactc ttccagattt tctcggactc cgcgcatcgc cgtaccactt
     1501 caaaacaccc aagcacagca ctgatgaatc ccctaatgat tttggtaaaa atcattaagt
     1561 taaggtggat acacatcttg tcatatgatc aaatggtttc gcgaaaaatc aataatcaga
     1621 caacaagatg tgcgaactcg atattttaca cgactctctt taccaattct gccccgaatt
     1681 acacttaaaa cgactcaaca gcttaacgtt ggcttgccac gcattacttg actgtaaaac
     1741 tctcactctt accgaacttg gccgtaacct gccaaccaaa gcgagaacaa aacataacat
     1801 caaacgaatc gaccgattgt taggtaatcg tcacctccac aaagagcgac tcgctgtata
     1861 ccgttggcat gctagcttta tctgttcggg caatacgatg cccattgtac ttgttgactg
     1921 gtctgatatt cgtgagcaaa aacgacttat ggtattgcga gcttcagtcg cactacacgg
     1981 tcgttctgtt actctttatg agaaagcgtt cccgctttca gagcaatgtt caaagaaagc
     2041 tcatgaccaa tttctagccg accttgcgag cattctaccg agtaacacca caccgctcat
     2101 tgtcagtgat gctggcttta aagtgccatg gtataaatcc gttgagaagc tgggttggta
     2161 ctggttaagt cgagtaagag gaaaagtaca atatgcagac ctaggagcgg aaaactggaa
     2221 acctatcagc aacttacatg atatgtcatc tagtcactca aagactttag gctataagag
     2281 gctgactaaa agcaatccaa tctcatgcca aattctattg tataaatctc gctctaaagg
     2341 ccgaaaaaat cagcgctcga cacggactca ttgtcaccac ccgtcaccta aaatctactc
     2401 agcgtcggca aaggagccat gggttctagc aactaactta cctgttgaaa ttcgaacacc
     2461 caaacaactt gttaatatct attcgaagcg aatgcagatt gaagaaacct tccgagactt
     2521 gaaaagtcct gcctacggac taggcctacg ccatagccga acgagcagct cagagcgttt
     2581 tgatatcatg ctgctaatcg ccctgatgct tcaactaaca tgttggcttg cgggcgttca
     2641 tgctcagaaa caaggttggg acaagcactt ccaggctaac acagtcagaa atcgaaacgt
     2701 actctcaaca gttcgcttag gcatggaagt tttgcggcat tctggctaca caataacaag
     2761 ggaagactta ctcgtggctg caaccctact agctcaaaat ttattcacac atggttacgc
     2821 tttggggaaa ttatgagggg atctctcaga gcacagcata ctaaatttcc cctctttctt
     2881 cctctagggt gtcgttaatt acccgtacta aaggtttgga aaagaaaaaa gagaccgcct
     2941 cgtttctttt tcttcgtcga aaaaggcaat aaaaattttt atcacgtttc tttttcttga
     3001 aaattttttt tttgattttt ttctctttcg atgacctccc attgatattt aagttaataa
     3061 acggtcttca atttctcaag tttcagtttc atttttcttg ttctattaca acttttttta
     3121 cttcttgctc attagaaaga aagcatagca atctaatcta agttttctag aactagtgga
     3181 tcccccggga aaaatggaca agaagtactc cattgggctc gatatcggca caaacagcgt
     3241 cggttgggcc gtcattacgg acgagtacaa ggtgccgagc aaaaaattca aagttctggg
     3301 caataccgat cgccacagca taaagaagaa cctcattggc gccctcctgt tcgactccgg
     3361 ggagacggcc gaagccacgc ggctcaaaag aacagcacgg cgcagatata cccgcagaaa
     3421 gaatcggatc tgctacctgc aggagatctt tagtaatgag atggctaagg tggatgactc
     3481 tttcttccat aggctggagg agtccttttt ggtggaggag gataaaaagc acgagcgcca
     3541 cccaatcttt ggcaatatcg tggacgaggt ggcgtaccat gaaaagtacc caaccatata
     3601 tcatctgagg aagaagcttg tagacagtac tgataaggct gacttgcggt tgatctatct
     3661 cgcgctggcg catatgatca aatttcgggg acacttcctc atcgaggggg acctgaaccc
     3721 agacaacagc gatgtcgaca aactctttat ccaactggtt cagacttaca atcagctttt
     3781 cgaagagaac ccgatcaacg catccggagt tgacgccaaa gcaatcctga gcgctaggct
     3841 gtccaaatcc cggcggctcg aaaacctcat cgcacagctc cctggggaga agaagaacgg
     3901 cctgtttggt aatcttatcg ccctgtcact cgggctgacc cccaacttta aatctaactt
     3961 cgacctggcc gaagatgcca agcttcaact gagcaaagac acctacgatg atgatctcga
     4021 caatctgctg gcccagatcg gcgaccagta cgcagacctt tttttggcgg caaagaacct
     4081 gtcagacgcc attctgctga gtgatattct gcgagtgaac acggagatca ccaaagctcc
     4141 gctgagcgct agtatgatca agcgctatga tgagcaccac caagacttga ctttgctgaa
     4201 ggcccttgtc agacagcaac tgcctgagaa gtacaaggaa attttcttcg atcagtctaa
     4261 aaatggctac gccggataca ttgacggcgg agcaagccag gaggaatttt acaaatttat
     4321 taagcccatc ttggaaaaaa tggacggcac cgaggagctg ctggtaaagc ttaacagaga
     4381 agatctgttg cgcaaacagc gcactttcga caatggaagc atcccccacc agattcacct
     4441 gggcgaactg cacgctatcc tcaggcggca agaggatttc tacccctttt tgaaagataa
     4501 cagggaaaag attgagaaaa tcctcacatt tcggataccc tactatgtag gccccctcgc
     4561 ccggggaaat tccagattcg cgtggatgac tcgcaaatca gaagagacca tcactccctg
     4621 gaacttcgag gaagtcgtgg ataagggggc ctctgcccag tccttcatcg aaaggatgac
     4681 taactttgat aaaaatctgc ctaacgaaaa ggtgcttcct aaacactctc tgctgtacga
     4741 gtacttcaca gtttataacg agctcaccaa ggtcaaatac gtcacagaag ggatgagaaa
     4801 gccagcattc ctgtctggag agcagaagaa agctatcgtg gacctcctct tcaagacgaa
     4861 ccggaaagtt accgtgaaac agctcaaaga agactatttc aaaaagattg aatgtttcga
     4921 ctctgttgaa atcagcggag tggaggatcg cttcaacgca tccctgggaa cgtatcacga
     4981 tctcctgaaa atcattaaag acaaggactt cctggacaat gaggagaacg aggacattct
     5041 tgaggacatt gtcctcaccc ttacgttgtt tgaagatagg gagatgattg aagaacgctt
     5101 gaaaacttac gctcatctct tcgacgacaa agtcatgaaa cagctcaaga ggcgccgata
     5161 tacaggatgg gggcggctgt caagaaaact gatcaatggg atccgagaca agcagagtgg
     5221 aaagacaatc ctggattttc ttaagtccga tggatttgcc aaccggaact tcatgcagtt
     5281 gatccatgat gactctctca cctttaagga ggacatccag aaagcacaag tttctggcca
     5341 gggggacagt cttcacgagc acatcgctaa tcttgcaggt agcccagcta tcaaaaaggg
     5401 aatactgcag accgttaagg tcgtggatga actcgtcaaa gtaatgggaa ggcataagcc
     5461 cgagaatatc gttatcgaga tggcccgaga gaaccaaact acccagaagg gacagaagaa
     5521 cagtagggaa aggatgaaga ggattgaaga gggtataaaa gaactggggt cccaaatcct
     5581 taaggaacac ccagttgaaa acacccagct tcagaatgag aagctctacc tgtactacct
     5641 gcagaacggc agggacatgt acgtggatca ggaactggac atcaatcggc tctccgacta
     5701 cgacgtggat catatcgtgc cccagtcttt tctcaaagat gattctattg ataataaagt
     5761 gttgacaaga tccgataaaa atagagggaa gagtgataac gtcccctcag aagaagttgt
     5821 caagaaaatg aaaaattatt ggcggcagct gctgaacgcc aaactgatca cacaacggaa
     5881 gttcgataat ctgactaagg ctgaacgagg tggcctgtct gagttggata aagccggctt
     5941 catcaaaagg cagcttgttg agacacgcca gatcaccaag cacgtggccc aaattctcga
     6001 ttcacgcatg aacaccaagt acgatgaaaa tgacaaactg attcgagagg tgaaagttat
     6061 tactctgaag tctaagctgg tctcagattt cagaaaggac tttcagtttt ataaggtgag
     6121 agagatcaac aattaccacc atgcgcatga tgcctacctg aatgcagtgg taggcactgc
     6181 acttatcaaa aaatatccca agcttgaatc tgaatttgtt tacggagact ataaagtgta
     6241 cgatgttagg aaaatgatcg caaagtctga gcaggaaata ggcaaggcca ccgctaagta
     6301 cttcttttac agcaatatta tgaatttttt caagaccgag attacactgg ccaatggaga
     6361 gattcggaag cgaccactta tcgaaacaaa cggagaaaca ggagaaatcg tgtgggacaa
     6421 gggtagggat ttcgcgacag tccggaaggt cctgtccatg ccgcaggtga acatcgttaa
     6481 aaagaccgaa gtacagaccg gaggcttctc caaggaaagt atcctcccga aaaggaacag
     6541 cgacaagctg atcgcacgca aaaaagattg ggaccccaag aaatacggcg gattcgattc
     6601 tcctacagtc gcttacagtg tactggttgt ggccaaagtg gagaaaggga agtctaaaaa
     6661 actcaaaagc gtcaaggaac tgctgggcat cacaatcatg gagcgatcaa gcttcgaaaa
     6721 aaaccccatc gactttctcg aggcgaaagg atataaagag gtcaaaaaag acctcatcat
     6781 taagcttccc aagtactctc tctttgagct tgaaaacggc cggaaacgaa tgctcgctag
     6841 tgcgggcgag ctgcagaaag gtaacgagct ggcactgccc tctaaatacg ttaatttctt
     6901 gtatctggcc agccactatg aaaagctcaa agggtctccc gaagataatg agcagaagca
     6961 gctgttcgtg gaacaacaca aacactacct tgatgagatc atcgagcaaa taagcgaatt
     7021 ctccaaaaga gtgatcctcg ccgacgctaa cctcgataag gtgctttctg cttacaataa
     7081 gcacagggat aagcccatca gggagcaggc agaaaacatt atccacttgt ttactctgac
     7141 caacttgggc gcgcctgcag ccttcaagta cttcgacacc accatagaca gaaagcggta
     7201 cacctctaca aaggaggtcc tggacgccac actgattcat cagtcaatta cggggctcta
     7261 tgaaacaaga atcgacctct ctcagctcgg tggagacagc agggctgacc ccaagaagaa
     7321 gaggaaggtg tgatctcttc tcgagtcatg taattagtta tgtcacgctt acattcacgc
     7381 cctcccccca catccgctct aaccgaaaag gaaggagtta gacaacctga agtctaggtc
     7441 cctatttatt tttttatagt tatgttagta ttaagaacgt tatttatatt tcaaattttt
     7501 cttttttttc tgtacagacg cgtgtacgca tgtaacatta tactgaaaac cttgcttgag
     7561 aaggttttgg gacgctcgaa ggctttaatt tgcggccggt acccaattcg ccctatagtg
     7621 agtcgtatta cgcgcgctca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     7681 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     7741 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggacgc
     7801 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     7861 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     7921 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     7981 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     8041 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     8101 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     8161 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     8221 gaattttaac aaaatattaa cgcttacaat ttcctgatgc ggtattttct ccttacgcat
     8281 ctgtgcggta tttcacaccg catagatcgg caagtgcaca aacaatactt aaataaatac
     8341 tactcagtaa taacctattt cttagcattt ttgacgaaat ttgctatttt gttagagtct
     8401 tttacaccat ttgtctccac acctccgctt acatcaacac caataacgcc atttaatcta
     8461 agcgcatcac caacattttc tggcgtcagt ccaccagcta acataaaatg taagctttcg
     8521 gggctctctt gccttccaac ccagtcagaa atcgagttcc aatccaaaag ttcacctgtc
     8581 ccacctgctt ctgaatcaaa caagggaata aacgaatgag gtttctgtga agctgcactg
     8641 agtagtatgt tgcagtcttt tggaaatacg agtcttttaa taactggcaa accgaggaac
     8701 tcttggtatt cttgccacga ctcatctcca tgcagttgga cgatatcaat gccgtaatca
     8761 ttgaccagag ccaaaacatc ctccttaggt tgattacgaa acacgccaac caagtatttc
     8821 ggagtgcctg aactattttt atatgctttt acaagacttg aaattttcct tgcaataacc
     8881 gggtcaattg ttctctttct attgggcaca catataatac ccagcaagtc agcatcggaa
     8941 tctagagcac attctgcggc ctctgtgctc tgcaagccgc aaactttcac caatggacca
     9001 gaactacctg tgaaattaat aacagacata ctccaagctg cctttgtgtg cttaatcacg
     9061 tatactcacg tgctcaatag tcaccaatgc cctccctctt ggccctctcc ttttcttttt
     9121 tcgaccgaat taattcttaa tcggcaaaaa aagaaaagct ccggatcaag attgtacgta
     9181 aggtgacaag ctatttttca ataaagaata tcttccacta ctgccatctg gcgtcataac
     9241 tgcaaagtac acatatatta cgatgctgtt ctattaaatg cttcctatat tatatatata
     9301 gtaatgtcgt tgatctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     9361 gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     9421 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac
     9481 cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta
     9541 atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg
     9601 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     9661 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     9721 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     9781 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     9841 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     9901 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     9961 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
    10021 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
    10081 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
    10141 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
    10201 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa
    10261 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
    10321 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
    10381 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
    10441 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
    10501 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
    10561 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
    10621 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
    10681 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
    10741 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
    10801 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
    10861 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
    10921 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
    10981 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
    11041 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
    11101 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
    11161 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
    11221 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
    11281 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
    11341 ttttacggtt cctggccttt tgctggcctt ttgctcgggg gatctgccaa ttgaacataa
    11401 catggtagtt acatatacta gtaatatggt tcggcacaca ttaaaagtat aaaaactatc
    11461 tgaattacga attacatata ttggtcataa aaatcaatca atcatcgtgt gttttatatg
    11521 tctcttatct aagtataaga atatccatag ttaatattca cttacgctac cttttaacct
    11581 gtaatcattg tcaacaggat atgttaacga cccacattga taaacgctag tatttctttt
    11641 tcctcttctt attggccggc tgtctctata ctcccctata gtctgtttct tttcgtttcg
    11701 attgtcccta tcagtgatag agatggcgca catggtacgc tgtggtgctc gcggctggga
    11761 acgaaactct gggagctgcg attggcagga ccatgcaata atctcctggt tttagagcta
    11821 gaaatagcaa gttaaaataa ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg
    11881 gtgctttttt ctcgagccat atccaacttc caatttaatc tttctttttt aattttcact
    11941 tatttgcgat acagaaagaa aaaagcgata gtaactattg aattttgttt ggatttggtt
    12001 agattagata tggtttctct ttatatttac atgctaaaaa tgggctacac cagagataca
    12061 taattagata tatatacgcc agtacacctt atcggcccaa gccttgtccc aaggcagcgt
    12121 tttgttcttg gaaacgctgc cctacacgtt cgctatgctt caagaacttt tctgagcact
    12181 tcatgatgca tgtttgttcc ttattggtta gctttgatgt tgtgaagtca ttgacacagt
    12241 ctgtgaaaca tctttctacc agattagagt acaaacgcat gaaatccttc atttgctttt
    12301 gttccactac tttttggaac tcttgttgtt ctttcctgca ggtagtcgtg caatgtatga
    12361 ctttaagatt tgtgagcagg aagaaaaggg agaatcttct aacgataaac ccttgaaaaa
    12421 ctgggtagac tacgctatgt tgagttgcta cgcaggctgc acaattacac gagaatgctc
    12481 ccgcctagga tttaaggcta agggacgtgc aatgcagacg acagatctaa atgaccgtgt
    12541 cggtgaagtg ttcgccaaac ttttcggtta acacatgcag tgatgcacgc gcgatggtgc
    12601 taagttacat atatatatat atgtatagcc atagtgatgt ctaagtaacc tttatggtat
    12661 atttcttaat gtggaaagat actagcgcgc gcacccacac acaagcttcg tcttttcttg
    12721 aagaaaagag gaagctcgct aaatgggatt ccactttccg ttccctgcca gctgatggaa
    12781 aaaggttagt ggaacgatga agaataaaaa gagagatcca ctgaggtgaa atttcagctg
    12841 acagcgagtt tcatgatcgt gatgaacaat ggtaacgagt tgtggctgtt gccagggagg
    12901 gtggttctca acttttaatg tatggccaaa tcgctacttg ggtttgttat ataacaaaga
    12961 agaaataatg aactgattct cttcctcctt cttgtccttt cttaattctg ttgtaattac
    13021 cttcctttgt aatttttttt gtaattattc ttcttaataa tccaaacaaa cacacatatt
    13081 acaatatatt aaa
//
