LOCUS       sequence-324490-       13833 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PphlF_A4NT, 1x cascade + [dCas9 + PA4-mVenus]. (Integration of 429; 
            pOSIP).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13833)
  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 13833)
  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: pJK434
FEATURES             Location/Qualifiers
     source          1..13833
                     /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(7019..7062)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     promoter        7083..7117
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     terminator      7861..7932
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      7948..7975
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_RNA        8072..8147
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        8442..8476
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     misc_RNA        8497..8572
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    8660..8680
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      8735..8762
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(8811..8842)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     protein_bind    8919..8952
                     /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(8959..9672)
                     /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             9761..9769
                     /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             9781..9803
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             9811..10821
                     /codon_start=1
                     /gene="int"
                     /product="integrase from phage 186"
                     /label=phage 186 integrase
                     /translation="MTVRKNPAGGWICELYPNGAKGKRIRKKFATKGEALAFEQYTVQN
                     PWQEEKEDRRTLKGLVDSWYSAHGITLKDGLKRQLAMHHAFECMGEPLARDFDAQMFSR
                     YREKRLKGEYARSNRVKEVSPRTLNLELAYFRAVFNELNRLGEWKGENPLKNMRPFRTE
                     EMEMTWLTHDQISQLLGECNRHDHPDLETVVRICLATGARWSEAESLRKSQLAKYKITY
                     TNTKGRKNRTVPISKELYESLPDDKKGRLFSDCYGAFRSALERTGIELPAGQLTHVLRH
                     TFASHFMMNGGNILVLQRVLGHTDIKMTMRYAHFAPDHLEDAVKLNPLVHITNSK"
     primer_bind     complement(10984..11003)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             10995..11810
                     /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(11068..11087)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      11932..12320
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     protein_bind    12331..12378
                     /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    12391..12605
                     /gene="attP"
                     /label=phage 186 attP
                     /bound_moiety="phage 186 integrase"
                     /note="attachment site of phage 186"
     terminator      complement(12722..12968)
                     /label=lambda tL3 terminator
                     /note="transcription terminator tL3 from phage lambda"
     primer_bind     complement(13059..13081)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     terminator      13291..13377
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      13474..13560
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      13657..13743
                     /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 gacaaattct agtgcttgga 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 tgcgtttata cctagtaata ggcgtatcac gaggcagaat
     7021 ttcagataaa aaaaatcctt agctttcgct aaggatgatt tctggaattc taaagatcta
     7081 cttttacagc tagctcagtc ctaggtatta tgctagcctg aagtacgtct gagcgtgata
     7141 cccgctcact gaagatgtcc cggtagggcc gaaacgtacc tctacaaata attttgttta
     7201 actatggact atgtttgaaa gggagaaata ctagatggca cgtaccccga gccgtagcag
     7261 cattggtagc ctgcgtagtc cgcataccca taaagcaatt ctgaccagca ccattgaaat
     7321 cctgaaagaa tgtggttata gcggtctgag cattgaaagc gttgcacgtc gtgccggtgc
     7381 aagcaaaccg accatttatc gttggtggac caataaagca gcactgattg ccgaagtgta
     7441 tgaaaatgaa agcgaacagg tgcgtaaatt tccggatctg ggtagcttta aagccgatct
     7501 ggattttctg ctgcgtaatc tgtggaaagt ttggcgtgaa accatttgtg gtgaagcatt
     7561 tcgttgtgtt attgcagaag cacagctgga ccctgcaacc ctgacccagc tgaaagatca
     7621 gtttatggaa cgtcgtcgtg agatgccgaa aaaactggtt gaaaatgcca ttagcaatgg
     7681 tgaactgccg aaagatacca atcgtgaact gctgctggat atgatttttg gtttttgttg
     7741 gtatcgcctg ctgaccgaac agctgaccgt tgaacaggat attgaagaat ttaccttcct
     7801 gctgattaat ggtgtttgtc cgggtacaca gcgttaacta gggcccatac ccccaggcat
     7861 caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg
     7921 gtgaacgctc tctactagag tcacactggc tcaccttcgg gtgggccttt ctgcgtttat
     7981 atacgcgacg tacggtggaa tctgattcgt taccaattga catgatacga aacgtaccgt
     8041 atcgttaagg tataatacca gctagttgtg ggttttagag ctagaaatag caagttaaaa
     8101 taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt ttttctcggt
     8161 accaaagacg aacaataaga cgctgaaaag cgtctttttt cgttttggtc cttcagccaa
     8221 aaaacttaag aagcttttta cacctccaca actagctggt attatgctag cataatacct
     8281 aggactgagc tgttttagag ctagaaatag caagttaaaa taaggctagt ccgattgcaa
     8341 cttgaaaaag tggcaccgag tcggtgcttt ttttctcggt accaaattcc agaaaagaga
     8401 cgctttcgag cgtctttttt cgttttggtc cgataggatc ctttacacct agctcagtcc
     8461 taggtattat gctagcataa tacccgtgtg acccgtgttt tagagctaga aatagcaagt
     8521 taaaataagg ctagtccgtt atcaacttga aaaagtggca ccgagtcggt gctttttttc
     8581 tcggtaccaa agacgaacaa taagacgctg aaaagcgtct tttttcgttt tggtccttta
     8641 tctagggtac gggttttgct actagtagcg gccgctgcag gcatgcctcg agatgcatgg
     8701 cgcctaacct aaactgacag gcatcaaatt aagcagaagg ccatcctgac ggatggcctt
     8761 tttgcgtttc gaacaattga aaaaacctcg cgccttacct gttgagtaat agtcaaaagc
     8821 ctccggtcgg aggcttttga ctttctgctt actgaatttc ggtggtgccg ttaattaacc
     8881 ggtgggccct catgataata atggtttctt agacgtccga agttcctatt ctctagaaag
     8941 tataggaact tccctaggtc agccaaacgt ctcttcaggc cactgactag cgataacttt
     9001 ccccacaacg gaacaactct cattgcatgg gatcattggg tactgtgggt ttagtggttg
     9061 taaaaacacc tgaccgctat ccctgatcag tttcttgaag gtaaactcat cacccccaag
     9121 tctggctatg cagaaatcac ctggctcaac agcctgctca gggtcaacga gaattaacat
     9181 tccgtcagga aagcttggct tggagcctgt tggtgcggtc atggaattac cttcaacctc
     9241 aagccagaat gcagaatcac tggctttttt ggttgtgctt acccatctct ccgcatcacc
     9301 tttggtaaag gttctaagct taggtgagaa catccctgcc tgaacatgag aaaaaacagg
     9361 gtactcatac tcacttctaa gtgacggctg catactaacc gcttcataca tctcgtagat
     9421 ttctctggcg attgaagggc taaattcttc aacgctaact ttgagaattt ttgtaagcaa
     9481 tgcggcgtta taagcattta atgcattgat gccattaaat aaagcaccaa cgcctgactg
     9541 ccccatcccc atcttgtctg cgacagattc ctgggataag ccaagttcat ttttcttttt
     9601 ttcataaatt gctttaaggc gacgtgcgtc ctcaagctgc tcttgtgtta atggtttctt
     9661 ttttgtgctc atacgttaaa tctatcaccg caagggataa atatctaaca ccgtgcgtgt
     9721 tgactatttt acctctggcg gtgataatgg ttgcatgtac taaggaggtt gtatacgcgt
     9781 tttgtttaac tttaagaagg agatatacat atgaccgtcc gtaaaaatcc ggctggcggt
     9841 tggatttgtg aactctaccc aaacggtgca aaaggcaaac gtatcagaaa gaaattcgct
     9901 actaaaggcg aggctctggc gtttgagcag tacaccgttc aaaacccgtg gcaggaagaa
     9961 aaggaagaca ggcgcacgtt aaaagggctg gttgattcat ggtatagcgc tcatggcatt
    10021 acactgaaag atggtttgaa acgccagtta gccatgcacc atgcttttga gtgtatgggc
    10081 gaaccactcg cacgcgattt cgatgcgcag atgttttccc gctaccgaga aaaacggtta
    10141 aaaggtgagt atgcccgttc aaacagagtg aaagaggtat cgcctcgcac gcttaatctt
    10201 gagctggcct acttccgggc ggtgttcaat gagctaaacc gcctcggaga atggaagggt
    10261 gaaaacccac taaaaaatat gcgcccattc cgcacagaag aaatggaaat gacttggcta
    10321 actcacgacc aaatttcgca actgctcgga gagtgtaacc gacatgacca ccctgattta
    10381 gaaaccgtgg taagaatctg tctcgcaact ggcgcacggt ggtctgaggc cgagagtctg
    10441 agaaaaagcc aactcgcgaa atacaaaatc acatacacca acacgaaagg tagaaaaaat
    10501 cgcaccgtcc caatcagcaa agagctctat gagtctctgc ctgatgataa aaagggccgg
    10561 ttgttcagtg attgttatgg cgcgttccgg tcagctttgg aaagaacagg tatcgaacta
    10621 ccggcaggac aacttaccca cgttttacgt cacaccttcg ctagtcactt tatgatgaat
    10681 ggcggtaata ttttggtgtt gcagcgcgta ctcggccata cagatataaa aatgaccatg
    10741 cgatatgcgc actttgcacc cgaccattta gaggatgcag ttaaactaaa tcctttagtt
    10801 cacattacta acagcaaata agttatggag gttgtatgtc agttcggcta gcccatgggt
    10861 atggacagtt ttccctttga tatgtaacgc acgttgtgtc tcaaaatctc tgatgttaca
    10921 ttgcacaaga taaaaatata tcatcatgaa caataaaact gtctgcttac ataaacagta
    10981 atacaagggg tgttatgagc catattcaac gggaaacgtc ttgctcccgt ccgcgcttaa
    11041 actccaacat ggacgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat
    11101 caggtgcgac aatctatcgc ttgtatggga agcccgatgc gccagagttg tttctgaaac
    11161 atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtccgtctc aactggctga
    11221 cggagtttat gcctctcccg accatcaagc attttatccg tactcctgat gatgcgtggt
    11281 tactcaccac cgcgattcct gggaaaacag ccttccaggt attagaagaa tatcctgatt
    11341 caggtgaaaa tattgttgat gcgctggccg tgttcctgcg ccggttacat tcgattcctg
    11401 tttgtaattg tccttttaac agcgatcgtg tatttcgtct tgctcaggcg caatcacgca
    11461 tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg
    11521 aacaagtctg gaaagaaatg cacaagctct tgccattctc accggattca gtcgtcactc
    11581 atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg
    11641 atgttggacg ggtcggaatc gcagaccgtt accaggacct tgccattctt tggaactgcc
    11701 tcggtgagtt ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc
    11761 ctgatatgaa taaattgcag tttcatttga tgctcgatga gtttttctaa taatactagc
    11821 agaaatcatc cttagcgaaa gctaaggatt ttttttatct gattaccgcc tttgagtgag
    11881 cgtcgaccta gtgcggccgc aagatccggc cacgatgcgt ccggcgtaga ggatctgaag
    11941 atcagcagtt caacctgttg atagtacgta ctaagctctc atgtttcacg tactaagctc
    12001 tcatgtttaa cgtactaagc tctcatgttt aacgaactaa accctcatgg ctaacgtact
    12061 aagctctcat ggctaacgta ctaagctctc atgtttcacg tactaagctc tcatgtttga
    12121 acaataaaat taatataaat cagcaactta aatagcctct aaggttttaa gttttataag
    12181 aaaaaaaaga atatataagg cttttaaagc ttttaaggtt taacggttgt ggacaacaag
    12241 ccagggatgt aacgcactga gaagccctta gagcctctca aagcaatttt gagtgacaca
    12301 ggaacactta acggctgaca tgggaattag gaagttccta ttccgaagtt cctattctct
    12361 agaaagtata ggaacttcca tatgccatgg gcgatggttc tgagtaacag ataatagaat
    12421 ggcgataaag tggcggtaga aatggcgaat aataggtaat tattggcaaa caatggcaat
    12481 ctatgtcaat gataaataac gcatactatt gattttcggt tgttcttgta ggaactcata
    12541 atcgcttggt cgctggttca agtccagcag gggccaccaa atttaagtag taaatacata
    12601 cagttaggcc actctctcga gtggcctttt tgttgcctcg ttttcgagtg gcgatagaat
    12661 ggcggtggat tttttaccgc cactttcctg aggagtgcat agctagcttc ttcgtctgtt
    12721 tctactggta ttggcacaaa cctgattcca atttgagcaa ggctatgtgc catctcgata
    12781 ctcgttctta actcaacaga agatgctttg tgcatacagc ccctcgttta ttatttatct
    12841 cctcagccag ccgctgtgct ttcagtggat ttcggataac agaaaggccg ggaaataccc
    12901 agcctcgctt tgtaacggag tagagacgaa agtgattgcg cctacccgga tattatcgtg
    12961 aggatgcgtc atcgccatta attcactgat cagtgataag ctgtcaaaca tgagaattga
    13021 tccggctgcc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
    13081 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
    13141 tcagcgggtg ttggcgggtg tcggggcgca gccatgaccc agtcacgtag cgatagcgga
    13201 gtgtatgctg cacatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct
    13261 tcaagaatta attcccaatt ccccaggcat caaataaaac gaaaggctca gtcgaaagac
    13321 tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg
    13381 ccgggagcgg atttgaacgt tgcgaagcaa cggcccggag ggtggcgggc aggacgcccg
    13441 ccataaactg ccaggaatta attccccagg catcaaataa aacgaaaggc tcagtcgaaa
    13501 gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag taggacaaat
    13561 ccgccgggag cggatttgaa cgttgcgaag caacggcccg gagggtggcg ggcaggacgc
    13621 ccgccataaa ctgccaggaa ttaattcccc aggcatcaaa taaaacgaaa ggctcagtcg
    13681 aaagactggg cctttcgttt tatctgttgt ttgtcggtga acgctctcct gagtaggaca
    13741 aatccgccgg gagcggattt gaacgttgcg aagcaacggc ccggagggtg gcgggcagga
    13801 cgcccgccat aaactgccag gaattgggga tcg
//
