LOCUS       sequence-301567-       12893 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  dCas9 oscillator v1 (dCRv1) + [dCas9 + PA4-mVenus]. (Integration of 
            412; pOSIP).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12893)
  AUTHORS   Kuo J, Yuan R, Sanchez C, Paulsson J, Silver PA
  TITLE     Toward a translationally independent RNA-based synthetic oscillator 
            using deactivated CRISPR-Cas.
  JOURNAL   Nucleic Acids Res. 2020 Aug 20;48(14):8165-8177. doi: 
            10.1093/nar/gkaa557.
  PUBMED    32609820
REFERENCE   2  (bases 1 to 12893)
  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"; doi: "10"; journalName: 
            "Nucleic Acids Res"; date: "2020-08-20- 20"; volume: "48"; issue: 
            "14"; pages: "8165-8177"
COMMENT     Sequence Label: pJK422
FEATURES             Location/Qualifiers
     source          1..12893
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        128..177
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     RBS             207..218
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             227..940
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTLGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             227..940
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTLGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(257..285)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     880..900
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      1001..1087
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      complement(1100..1194)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(1215..1874)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1789..1808
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(1875..1977)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             complement(2007..2633)
                     /codon_start=1
                     /gene="tetR"
                     /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="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     2538..2557
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        2649..2704
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    2685..2703
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             2722..2733
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             join(2740..2766,2767..2769,2770..5256,5257..5259,
                     5260..6846)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     terminator      6870..6941
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6957..6984
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(7018..7061)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_RNA        7144..7219
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        7503..7537
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     misc_RNA        7558..7633
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    7721..7741
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      7796..7823
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(7872..7903)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     protein_bind    7980..8013
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             complement(8020..8733)
                     /codon_start=1
                     /gene="cIts"
                     /product="temperature-sensitive variant of the phage lambda
                     repressor"
                     /label=lambda repressor (ts)
                     /note="thermosensitivity is conferred by the A67T mutation"
                     /translation="MSTKKKPLTQEQLEDARRLKAIYEKKKNELGLSQESVADKMGMGQ
                     SGVGALFNGINALNAYNAALLTKILKVSVEEFSPSIAREIYEMYEAVSMQPSLRSEYEY
                     PVFSHVQAGMFSPKLRTFTKGDAERWVSTTKKASDSAFWLEVEGNSMTAPTGSKPSFPD
                     GMLILVDPEQAVEPGDFCIARLGGDEFTFKKLIRDSGQVFLQPLNPQYPMIPCNESCSV
                     VGKVIASQWPEETFG"
     RBS             8822..8830
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     RBS             8842..8864
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             8872..9882
                     /codon_start=1
                     /gene="int"
                     /product="integrase from phage 186"
                     /label=phage 186 integrase
                     /translation="MTVRKNPAGGWICELYPNGAKGKRIRKKFATKGEALAFEQYTVQN
                     PWQEEKEDRRTLKELVDSWYSAHGITLKDGLKRQLAMHHAFECMGEPLARDFDAQMFSR
                     YREKRLKGEYARSNRVKEVSPRTLNLELAYFRAVFNELNRLGEWKGENPLKNMRPFRTE
                     EMEMTWLTHDQISQLLGECNRHDHPDLETVVRICLATGARWSEAESLRKSQLAKYKITY
                     TNTKGRKNRTVPISKELYESLPDDKKGRLFSDCYGAFRSALERTGIELPAGQLTHVLRH
                     TFASHFMMNGGNILVLQRVLGHTDIKMTMRYAHFAPDHLEDAVKLNPLVHITNSK"
     primer_bind     complement(10045..10064)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             10056..10871
                     /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"
     primer_bind     complement(10129..10148)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      10993..11381
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     protein_bind    11392..11439
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     protein_bind    11452..11666
                     /gene="attP"
                     /label=phage 186 attP
                     /bound_moiety="phage 186 integrase"
                     /note="attachment site of phage 186"
     terminator      complement(11783..12029)
                     /label=lambda tL3 terminator
                     /note="transcription terminator tL3 from phage lambda"
     primer_bind     complement(12119..12141)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     terminator      12351..12437
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      12534..12620
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      12717..12803
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gaattcgagc tcggtacccg gggatcctct agagggctta ctaaaagcca gataacagta
       61 tgcgtatttg cgcgcctagt atgcgtctcc tttacacctc cacaactagc tggtattatg
      121 ctagcagctg tcaccggatg tgctttccgg tctgatgagt ccgtgaggac gaaacagcct
      181 ctacaaataa ttttgtttaa ctttacaaag aggagaaagg taccgcatga gtaaaggaga
      241 agaacttttc actggagttg tcccaattct tgttgaatta gatggtgatg ttaatgggca
      301 caaattttct gtcagtggag agggtgaagg tgatgcaaca tacggaaaac ttacccttaa
      361 attgatttgc actactggaa aactacctgt tccatggcca acacttgtca ctactttggg
      421 ttatggtgtt caatgctttg cgagataccc agatcatatg aaacagcatg actttttcaa
      481 gagtgccatg cccgaaggtt atgtacagga aagaactata tttttcaaag atgacgggaa
      541 ctacaagaca cgtgctgaag tcaagtttga aggtgatacc cttgttaata gaatcgagtt
      601 aaaaggtatt gattttaaag aagatggaaa cattcttgga cacaaattgg aatacaacta
      661 taactcacac aatgtataca tcacggcaga caaacaaaag aatggaatca aagcgaactt
      721 caaaattaga cacaacattg aagatggagg tgttcaacta gcagaccatt atcaacaaaa
      781 tactccaatt ggcgatggcc ctgtcctttt accagacaac cattacctgt cctaccaatc
      841 taagctttcg aaagatccca acgaaaagag agaccacatg gtccttcttg agtttgtaac
      901 agctgctggg attacacatg gcatggatga actatacaaa taagcttgat atcgaattcc
      961 tgcagcccgg gggatcccat tgtacgcgtg ctagaggcat caaataaaac gaaaggctca
     1021 gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag
     1081 gacaaatact agagcttgga ttctcaccaa taaaaaacgc ccggcggcaa ccgagcgttc
     1141 tgaacaaatc cagatggagt tctgaggtca ttactggatc tatcaacagg agtccaagcg
     1201 agctcgatat caaattacgc cccgccctgc cactcatcgc agtactgttg taattcatta
     1261 agcattctgc cgacatggaa gccatcacaa acggcatgat gaacctgaat cgccagcggc
     1321 atcagcacct tgtcgccttg cgtataatat ttgcccatgg tgaaaacggg ggcgaagaag
     1381 ttgtccatat tggccacgtt taaatcaaaa ctggtgaaac tcacccaggg attggctgag
     1441 acgaaaaaca tattctcaat aaacccttta gggaaatagg ccaggttttc accgtaacac
     1501 gccacatctt gcgaatatat gtgtagaaac tgccggaaat cgtcgtggta ttcactccag
     1561 agcgatgaaa acgtttcagt ttgctcatgg aaaacggtgt aacaagggtg aacactatcc
     1621 catatcacca gctcaccgtc tttcattgcc atacgaaatt ccggatgagc attcatcagg
     1681 cgggcaagaa tgtgaataaa ggccggataa aacttgtgct tatttttctt tacggtcttt
     1741 aaaaaggccg taatatccag ctgaacggtc tggttatagg tacattgagc aactgactga
     1801 aatgcctcaa aatgttcttt acgatgccat tgggatatat caacggtggt atatccagtg
     1861 atttttttct ccattttagc ttccttagct cctgaaaatc tcgataactc aaaaaatacg
     1921 cccggtagtg atcttatttc attatggtga aagttggaac ctcttacgtg ccgatcaacg
     1981 tctcattttc gccagatatc gacgtcttaa gacccacttt cacatttaag ttgtttttct
     2041 aatccgcata tgatcaattc aaggccgaat aagaaggctg gctctgcacc ttggtgatca
     2101 aataattcga tagcttgtcg taataatggc ggcatactat cagtagtagg tgtttccctt
     2161 tcttctttag cgacttgatg ctcttgatct tccaatacgc aacctaaagt aaaatgcccc
     2221 acagcgctga gtgcatataa tgcattctct agtgaaaaac cttgttggca taaaaaggct
     2281 aattgatttt cgagagtttc atactgtttt tctgtaggcc gtgtacctaa atgtactttt
     2341 gctccatcgc gatgacttag taaagcacat ctaaaacttt tagcgttatt acgtaaaaaa
     2401 tcttgccagc tttccccttc taaagggcaa aagtgagtat ggtgcctatc taacatctca
     2461 atggctaagg cgtcgagcaa agcccgctta ttttttacat gccaatacaa tgtaggctgc
     2521 tctacaccta gcttctgggc gagtttacgg gttgttaaac cttcgattcc gacctcatta
     2581 agcagctcta atgcgctgtt aatcacttta cttttatcta atctagacat cattaattcc
     2641 taatttttgt tgacactcta tcgttgatag agttatttta ccactcccta tcagtgatag
     2701 agaaaagaat tcaaaagatc taaagaggag aaaggatcta tggataagaa atactcaata
     2761 ggcttagcta tcggcacaaa tagcgtcgga tgggcggtga tcactgatga atataaggtt
     2821 ccgtctaaaa agttcaaggt tctgggaaat acagaccgcc acagtatcaa aaaaaatctt
     2881 ataggggctc ttttatttga cagtggagag acagcggaag cgactcgtct caaacggaca
     2941 gctcgtagaa ggtatacacg tcggaagaat cgtatttgtt atctacagga gattttttca
     3001 aatgagatgg cgaaagtaga tgatagtttc tttcatcgac ttgaagagtc ttttttggtg
     3061 gaagaagaca agaagcatga acgtcatcct atttttggaa atatagtaga tgaagttgct
     3121 tatcatgaga aatatccaac tatctatcat ctgcgaaaaa aattggtaga ttctactgat
     3181 aaagcggatt tgcgcttaat ctatttggcc ttagcgcata tgattaagtt tcgtggtcat
     3241 tttttgattg agggagattt aaatcctgat aatagtgatg tggacaaact atttatccag
     3301 ttggtacaaa cctacaatca attatttgaa gaaaacccta ttaacgcaag tggagtagat
     3361 gctaaagcga ttctttctgc acgattgagt aaatcaagac gattagaaaa tctcattgct
     3421 cagctccccg gtgagaagaa aaatggctta tttgggaatc tcattgcttt gtcattgggt
     3481 ttgaccccta attttaaatc aaattttgat ttggcagaag atgctaaatt acagctttca
     3541 aaagatactt acgatgatga tttagataat ttattggcgc aaattggaga tcaatatgct
     3601 gatttgtttt tggcagctaa gaatttatca gatgctattt tactttcaga tatcctaaga
     3661 gtaaatactg aaataactaa ggctccccta tcagcttcaa tgattaaacg ctacgatgaa
     3721 catcatcaag acttgactct tttaaaagct ttagttcgac aacaacttcc agaaaagtat
     3781 aaagaaatct tttttgatca atcaaaaaac ggatatgcag gttatattga tgggggagct
     3841 agccaagaag aattttataa atttatcaaa ccaattttag aaaaaatgga tggtactgag
     3901 gaattattgg tgaaactaaa tcgtgaagat ttgctgcgca agcaacggac ctttgacaac
     3961 ggctctattc cccatcaaat tcacttgggt gagctgcatg ctattttgag aagacaagaa
     4021 gacttttatc catttttaaa agacaatcgt gagaagattg aaaaaatctt gacttttcga
     4081 attccttatt atgttggtcc attggcgcgt ggcaatagtc gttttgcatg gatgactcgg
     4141 aagtctgaag aaacaattac cccatggaat tttgaagaag ttgtcgataa aggtgcttca
     4201 gctcaatcat ttattgaacg catgacaaac tttgataaaa atcttccaaa tgaaaaagta
     4261 ctaccaaaac atagtttgct ttatgagtat tttacggttt ataacgaatt gacaaaggtc
     4321 aaatatgtta ctgaaggaat gcgaaaacca gcatttcttt caggtgaaca gaagaaagcc
     4381 attgttgatt tactcttcaa aacaaatcga aaagtaaccg ttaagcaatt aaaagaagat
     4441 tatttcaaaa aaatagaatg ttttgatagt gttgaaattt caggagttga agatagattt
     4501 aatgcttcat taggtaccta ccatgatttg ctaaaaatta ttaaagataa agattttttg
     4561 gataatgaag aaaatgaaga tatcttagag gatattgttt taacattgac cttatttgaa
     4621 gatagggaga tgattgagga aagacttaaa acatatgctc acctctttga tgataaggtg
     4681 atgaaacagc ttaaacgtcg ccgttatact ggttggggac gtttgtctcg aaaattgatt
     4741 aatggtatta gggataagca atctggcaaa acaatattag attttttgaa atcagatggt
     4801 tttgccaatc gcaattttat gcagctgatc catgatgata gtttgacatt taaagaagac
     4861 attcaaaaag cacaagtgtc tggacaaggc gatagtttac atgaacatat tgcaaattta
     4921 gctggtagcc ctgctattaa aaaaggtatt ttacagactg taaaagttgt tgatgaattg
     4981 gtcaaagtaa tggggcggca taagccagaa aatatcgtta ttgaaatggc acgtgaaaat
     5041 cagacaactc aaaagggcca gaaaaattcg cgagagcgta tgaaacgaat cgaagaaggt
     5101 atcaaagaat taggaagtca gattcttaaa gagcatcctg ttgaaaatac tcaattgcaa
     5161 aatgaaaagc tctatctcta ttatctccaa aatggaagag acatgtatgt ggaccaagaa
     5221 ttagatatta atcgtttaag tgattatgat gtcgatgcca ttgttccaca aagtttcctt
     5281 aaagacgatt caatagacaa taaggtctta acgcgttctg ataaaaatcg tggtaaatcg
     5341 gataacgttc caagtgaaga agtagtcaaa aagatgaaaa actattggag acaacttcta
     5401 aacgccaagt taatcactca acgtaagttt gataatttaa cgaaagctga acgtggaggt
     5461 ttgagtgaac ttgataaagc tggttttatc aaacgccaat tggttgaaac tcgccaaatc
     5521 actaagcatg tggcacaaat tttggatagt cgcatgaata ctaaatacga tgaaaatgat
     5581 aaacttattc gagaggttaa agtgattacc ttaaaatcta aattagtttc tgacttccga
     5641 aaagatttcc aattctataa agtacgtgag attaacaatt accatcatgc ccatgatgcg
     5701 tatctaaatg ccgtcgttgg aactgctttg attaagaaat atccaaaact tgaatcggag
     5761 tttgtctatg gtgattataa agtttatgat gttcgtaaaa tgattgctaa gtctgagcaa
     5821 gaaataggca aagcaaccgc aaaatatttc ttttactcta atatcatgaa cttcttcaaa
     5881 acagaaatta cacttgcaaa tggagagatt cgcaaacgcc ctctaatcga aactaatggg
     5941 gaaactggag aaattgtctg ggataaaggg cgagattttg ccacagtgcg caaagtattg
     6001 tccatgcccc aagtcaatat tgtcaagaaa acagaagtac agacaggcgg attctccaag
     6061 gagtcaattt taccaaaaag aaattcggac aagcttattg ctcgtaaaaa agactgggat
     6121 ccaaaaaaat atggtggttt tgatagtcca acggtagctt attcagtcct agtggttgct
     6181 aaggtggaaa aagggaaatc gaagaagtta aaatccgtta aagagttact agggatcaca
     6241 attatggaaa gaagttcctt tgaaaaaaat ccgattgact ttttagaagc taaaggatat
     6301 aaggaagtta aaaaagactt aatcattaaa ctacctaaat atagtctttt tgagttagaa
     6361 aacggtcgta aacggatgct ggctagtgcc ggagaattac aaaaaggaaa tgagctggct
     6421 ctgccaagca aatatgtgaa ttttttatat ttagctagtc attatgaaaa gttgaagggt
     6481 agtccagaag ataacgaaca aaaacaattg tttgtggagc agcataagca ttatttagat
     6541 gagattattg agcaaatcag tgaattttct aagcgtgtta ttttagcaga tgccaattta
     6601 gataaagttc ttagtgcata taacaaacat agagacaaac caatacgtga acaagcagaa
     6661 aatattattc atttatttac gttgacgaat cttggagctc ccgctgcttt taaatatttt
     6721 gatacaacaa ttgatcgtaa acgatatacg tctacaaaag aagttttaga tgccactctt
     6781 atccatcaat ccatcactgg tctttatgaa acacgcattg atttgagtca gctaggaggt
     6841 gactaactcg agtaaggatc tccaggcatc aaataaaacg aaaggctcag tcgaaagact
     6901 gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct ctactagagt cacactggct
     6961 caccttcggg tgggcctttc tgcgtttatc ctagtaatag gcgtatcacg aggcagaatt
     7021 tcagataaaa aaaatcctta gctttcgcta aggatgattt ctggaattct aaagatctca
     7081 gaaatcattt tacaccaacg ggtcacacgg gtattatgct agcataatac cagctagttg
     7141 tgggttttag agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa
     7201 agtggcaccg agtcggtgct ttttttctcg gtaccaaatt ccagaaaaga ggcctcccga
     7261 aaggggggcc ttttttcgtt ttggtccgct taagcttttt acacctccac aactagctgg
     7321 tattatgcta gcataatacc taggactgag ctgttttaga gctagaaata gcaagttaaa
     7381 ataaggctag tccgattgca acttgaaaaa gtggcaccga gtcggtgctt tttttctcgg
     7441 taccaaattc cagaaaagag acgctttcga gcgtcttttt tcgttttggt ccgataggat
     7501 cctttacacc tagctcagtc ctaggtatta tgctagcata atacccgtgt gacccgtgtt
     7561 ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtggc
     7621 accgagtcgg tgcttttttt ctcggtacca aagacgaaca ataagacgct gaaaagcgtc
     7681 ttttttcgtt ttggtccttt atctagggta cgggttttgc tactagtagc ggccgctgca
     7741 ggcatgcctc gagatgcatg gcgcctaacc taaactgaca ggcatcaaat taagcagaag
     7801 gccatcctga cggatggcct ttttgcgttt cgaacaattg aaaaaacctc gcgccttacc
     7861 tgttgagtaa tagtcaaaag cctccggtcg gaggcttttg actttctgct tactgaattt
     7921 cggtggtgcc gttaattaac cggtgggccc tcatgataat aatggtttct tagacgtccg
     7981 aagttcctat tctctagaaa gtataggaac ttccctaggt cagccaaacg tctcttcagg
     8041 ccactgacta gcgataactt tccccacaac ggaacaactc tcattgcatg ggatcattgg
     8101 gtactgtggg tttagtggtt gtaaaaacac ctgaccgcta tccctgatca gtttcttgaa
     8161 ggtaaactca tcacccccaa gtctggctat gcagaaatca cctggctcaa cagcctgctc
     8221 agggtcaacg agaattaaca ttccgtcagg aaagcttggc ttggagcctg ttggtgcggt
     8281 catggaatta ccttcaacct caagccagaa tgcagaatca ctggcttttt tggttgtgct
     8341 tacccatctc tccgcatcac ctttggtaaa ggttctaagc ttaggtgaga acatccctgc
     8401 ctgaacatga gaaaaaacag ggtactcata ctcacttcta agtgacggct gcatactaac
     8461 cgcttcatac atctcgtaga tttctctggc gattgaaggg ctaaattctt caacgctaac
     8521 tttgagaatt tttgtaagca atgcggcgtt ataagcattt aatgcattga tgccattaaa
     8581 taaagcacca acgcctgact gccccatccc catcttgtct gcgacagatt cctgggataa
     8641 gccaagttca tttttctttt tttcataaat tgctttaagg cgacgtgcgt cctcaagctg
     8701 ctcttgtgtt aatggtttct tttttgtgct catacgttaa atctatcacc gcaagggata
     8761 aatatctaac accgtgcgtg ttgactattt tacctctggc ggtgataatg gttgcatgta
     8821 ctaaggaggt tgtatacgcg ttttgtttaa ctttaagaag gagatataca tatgaccgtc
     8881 cgtaaaaatc cggctggcgg ttggatttgt gaactctacc caaacggtgc aaaaggcaaa
     8941 cgtatcagaa agaaattcgc tactaaaggc gaggctctgg cgtttgagca gtacaccgtt
     9001 caaaacccgt ggcaggaaga aaaggaagac aggcgcacgt taaaagagct ggttgattca
     9061 tggtatagcg ctcatggcat tacactgaaa gatggtttga aacgccagtt agccatgcac
     9121 catgcttttg agtgtatggg cgaaccactc gcacgcgatt tcgatgcgca gatgttttcc
     9181 cgctaccgag aaaaacggtt aaaaggtgag tatgcccgtt caaacagagt gaaagaggta
     9241 tcgcctcgca cgcttaatct tgagctggcc tacttccggg cggtgttcaa tgagctaaac
     9301 cgcctcggag aatggaaggg tgaaaaccca ctaaaaaata tgcgcccatt ccgcacagaa
     9361 gaaatggaaa tgacttggct aactcacgac caaatttcgc aactgctcgg agagtgtaac
     9421 cgacatgacc accctgattt agaaaccgtg gtaagaatct gtctcgcaac tggcgcacgg
     9481 tggtctgagg ccgagagtct gagaaaaagc caactcgcga aatacaaaat cacatacacc
     9541 aacacgaaag gtagaaaaaa tcgcaccgtc ccaatcagca aagagctcta tgagtctctg
     9601 cctgatgata aaaagggccg gttgttcagt gattgttatg gcgcgttccg gtcagctttg
     9661 gaaagaacag gtatcgaact accggcagga caacttaccc acgttttacg tcacaccttc
     9721 gctagtcact ttatgatgaa tggcggtaat attttggtgt tgcagcgcgt actcggccat
     9781 acagatataa aaatgaccat gcgatatgcg cactttgcac ccgaccattt agaggatgca
     9841 gttaaactaa atcctttagt tcacattact aacagcaaat aagttatgga ggttgtatgt
     9901 cagttcggct agcccatggg tatggacagt tttccctttg atatgtaacg cacgttgtgt
     9961 ctcaaaatct ctgatgttac attgcacaag ataaaaatat atcatcatga acaataaaac
    10021 tgtctgctta cataaacagt aatacaaggg gtgttatgag ccatattcaa cgggaaacgt
    10081 cttgctcccg tccgcgctta aactccaaca tggacgctga tttatatggg tataaatggg
    10141 ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg cttgtatggg aagcccgatg
    10201 cgccagagtt gtttctgaaa catggcaaag gtagcgttgc caatgatgtt acagatgaga
    10261 tggtccgtct caactggctg acggagttta tgcctctccc gaccatcaag cattttatcc
    10321 gtactcctga tgatgcgtgg ttactcacca ccgcgattcc tgggaaaaca gccttccagg
    10381 tattagaaga atatcctgat tcaggtgaaa atattgttga tgcgctggcc gtgttcctgc
    10441 gccggttaca ttcgattcct gtttgtaatt gtccttttaa cagcgatcgt gtatttcgtc
    10501 ttgctcaggc gcaatcacgc atgaataacg gtttggttga tgcgagtgat tttgatgacg
    10561 agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat gcacaagctc ttgccattct
    10621 caccggattc agtcgtcact catggtgatt tctcacttga taaccttatt tttgacgagg
    10681 ggaaattaat aggttgtatt gatgttggac gggtcggaat cgcagaccgt taccaggacc
    10741 ttgccattct ttggaactgc ctcggtgagt tttctccttc attacagaaa cggctttttc
    10801 aaaaatatgg tattgataat cctgatatga ataaattgca gtttcatttg atgctcgatg
    10861 agtttttcta ataatactag cagaaatcat ccttagcgaa agctaaggat tttttttatc
    10921 tgattaccgc ctttgagtga gcgtcgacct agtgcggccg caagatccgg ccacgatgcg
    10981 tccggcgtag aggatctgaa gatcagcagt tcaacctgtt gatagtacgt actaagctct
    11041 catgtttcac gtactaagct ctcatgttta acgtactaag ctctcatgtt taacgaacta
    11101 aaccctcatg gctaacgtac taagctctca tggctaacgt actaagctct catgtttcac
    11161 gtactaagct ctcatgtttg aacaataaaa ttaatataaa tcagcaactt aaatagcctc
    11221 taaggtttta agttttataa gaaaaaaaag aatatataag gcttttaaag cttttaaggt
    11281 ttaacggttg tggacaacaa gccagggatg taacgcactg agaagccctt agagcctctc
    11341 aaagcaattt tgagtgacac aggaacactt aacggctgac atgggaatta ggaagttcct
    11401 attccgaagt tcctattctc tagaaagtat aggaacttcc atatgccatg ggcgatggtt
    11461 ctgagtaaca gataatagaa tggcgataaa gtggcggtag aaatggcgaa taataggtaa
    11521 ttattggcaa acaatggcaa tctatgtcaa tgataaataa cgcatactat tgattttcgg
    11581 ttgttcttgt aggaactcat aatcgcttgg tcgctggttc aagtccagca ggggccacca
    11641 aatttaagta gtaaatacat acagttaggc cactctctcg agtggccttt ttgttgcctc
    11701 gttttcgagt ggcgatagaa tggcggtgga ttttttaccg ccactttcct gaggagtgca
    11761 tagctagctt cttcgtctgt ttctactggt attggcacaa acctgattcc aatttgagca
    11821 aggctatgtg ccatctcgat actcgttctt aactcaacag aagatgcttt gtgcatacag
    11881 cccctcgttt attatttatc tcctcagcca gccgctgtgc tttcagtgga tttcggataa
    11941 cagaaaggcc gggaaatacc cagcctcgct ttgtaacgga gtagagacga aagtgattgc
    12001 gcctacccgg atattatcgt gaggatgcgt catcgccatt aattcactga tcagtgatag
    12061 ctgtcaaaca tgagaattga tccggctgcc tcgcgcgttt cggtgatgac ggtgaaaacc
    12121 tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca
    12181 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggcgca gccatgaccc
    12241 agtcacgtag cgatagcgga gtgtatgctg cacatgacat taacctataa aaataggcgt
    12301 atcacgaggc cctttcgtct tcaagaatta attcccaatt ccccaggcat caaataaaac
    12361 gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc
    12421 tcctgagtag gacaaatccg ccgggagcgg atttgaacgt tgcgaagcaa cggcccggag
    12481 ggtggcgggc aggacgcccg ccataaactg ccaggaatta attccccagg catcaaataa
    12541 aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg
    12601 ctctcctgag taggacaaat ccgccgggag cggatttgaa cgttgcgaag caacggcccg
    12661 gagggtggcg ggcaggacgc ccgccataaa ctgccaggaa ttaattcccc aggcatcaaa
    12721 taaaacgaaa ggctcagtcg aaagactggg cctttcgttt tatctgttgt ttgtcggtga
    12781 acgctctcct gagtaggaca aatccgccgg gagcggattt gaacgttgcg aagcaacggc
    12841 ccggagggtg gcgggcagga cgcccgccat aaactgccag gaattgggga tcg
//
