.) 1990 Oxford University Press
4924 Nucleic Acids Research, Vol. 18, No. 16
Nucleotide sequence of Clostridium botulinum C1 neurotoxin Daniel Hauser, Melvin W.Eklund2, Hisao Kurazono3, Thomas Binz3, Heiner Niemann 3, D.Michael Gill4, Patrice Boquet1 and Michel R.Popoff* Unite des Anaerobies and 1Unit' des Antigenes Bacteriens, Institut Pasteur, 28 rue du Dr. Roux-75724 Paris cedex 15, France, 2Northwest and Alaska Fisheries Center, 2725 Montlake Boulevard East, Seattle, WA 98112, USA, 3Institut fur Mikrobiologie an der Bundesforschungssanstalt fOr Viruskrankheiten der Tiere, Postfach 1149, 7400 TObingen, FRG and 4Department of Molecular Biology and Microbiology, Tufts University Schools of Medicine, Boston, MA 02111, USA EMBL accession no. X53751
Submitted July 13, 1990
Clostridiu,n botulinum neurotoxins consist of seven serologically distinct types: A, B, C1, D, E, F, and G. All of these are synthesized as single-chain polypeptides of about 150 kDa which are generally nicked after secretion to yield N-terminal light ( = 50 kDa) and C-terminal heavy (= 100 kDa) chains connected by a single disulfide bond. C. botulinum C I neurotoxin is encoded by a bacteriophage (1, 2). C. botulinum Cl neurotoxin DNA sequence shown below was derived from cloned C phage DNA fragments the sizes of which were less than 2.0 kb. The initial clone containing a Ci.neurotoxin coding DNA was identified using a mixed 29-mer probe [5 '-GA(T/C)CCIGTIGA(T/C)AA(T/C)AA(A/G)AA(T/C)ATICTITA-3'] which corresponds to Asp'2 to Tyr21 of the partial amino acid sequence published previously (3). Potential biohazards associated with the experiments described had been approved by the French National Control Committee. The complete C I DNA sequence shows a single open reading frame which begins with an ATG at position 207 and encodes for 1291 amino acids (MR 148,698 Da). Before the ATG a ShineDalgarno sequence can be localized from position 191 to 196. Cys437 and Cys453 are probably involved in a disulfide bridge linking the light and heavy chains. A candidate transcription terminator (AG= -6.2 kCal.mol-1) is underlined. C. botulinum Cl neurotoxin exhibits no similarity with exoenzyme I GAATCAATAG TAAGTCAAAG ATAATGCAAA 61 CT67TTGTAT AACATTTTCA TATAAT6ATA 121 TTATAATTGG ATG6TAT6TA ATGACAATAA 181 TTAGAAAGTT AGGAGATGTT AGTATTATGC 241 ATCCTGTTGA TAATAAAAAT ATTTTA7TATT 301 AGCCTGAAAA AGCCTTTCGC ATTACAGGAA 361 GAAATTCTAA TCCAAATTTA AATAAACCTC 421 ATGATCCTAA TTATTTGT ACTGATTCTG
481 A4TTATTTAA AA4AATTAAT 541 CAGATATACC CTTTCCTGGG
7TATATGGGT AATACCT6AT AGATTTTC6 A
CTCGAGTTAC AAGCCCTAAA AGTGGTTATT ACAAAGATCC ATTTTTAAAA GAAATTATAA TCTAGAGA4A TAG6AGAAGA ATTAATATAT AGACTTTCGA AATAACAATA CTCCAATT'A T6CTTTTGAT TTTGATGTA6 AAAACTA6AC AAGGTAACAA CT68TTAA4 ACT6GTA6CA ACTGGACCTA G4GAAAACAT TATAGATCCA GAAACTTCTA ACTTTTGC6G 7ACAAGA4GG ATTTGT76CT TTATCAATAA ATGCTAACAT ATAGTAATGC 7ACTAAT6AT GTA46G6 TTTTCATG66 ATCCAATACT AaTTTTAAT6 CATGAACTTA TATG847A8G CTATACCAAA TGATCAAACA ATTTCATCT6 TCTCAATATA AT7TGA6ATT AG6ATATGCA 6AAATATATG
601 ATTTTAACA66 TTTGATGTT 661 TAAATCCTAG T6TTATAATA 721 C8TTTAAATT AACTAACAAT 781 TTTCAATATC ACCTAGATTT 841 6TAGATTT7C TAAGTCTGAA 901 ATCATGCAAT 6CATAATTTA 961 TAACTAGTAA TATTTTTTAT 1021 CATTT7766 TCCAACTATA G4CCTTATTC 1081 A4 766ATTG6A TATTATAGA TCTATAGCTA 1141 CTTCAAGCTT T7ATAAAAT4AT7A466844AT 1201 TC7TA6TA6A ATCTTCAGGT TTACAG 1261 ATAA4CTTAC ACAAATATTT ACA766TTTA 1321 B6AAaATA6 A TCTTTCAAAT 6TATATACTC 1361 TtTAT7A6AT ACAAAAT76 TTtAATATAC 1441 6TCAA4AT4 T ATCTC6AAAT CCA4CATTAA *
AAGATTACAG TTAATACTGA TTATTTA6AC AATATT7TCT CTATC4CTTA GAGATGGAGA CAAG6TGCTA AAGGTGCACA TTTGTGGATA CAATAACAAT TAACAACTTT AATTATTCAG TAGATACTCA TTTAAATACA CTAGCTAATG
CTAAA76TGC AA6BAAATAT TTTGA7664
AAAGACTTAA TAGTATAACT ACTGCAAATC ATA6 7AGAA ACTTATTA6A AA6TATA64T
TAAATCGTAA TAA6TTTGTT 946TTAT6TA ACTACGCTAA AATATATAAT 6TACAA4ATA C6GTT6C6GC 8AATATATTA 64CGATAAT6
CTAAAA6TAA TTTAAAT7TA CTATTTAT76
GAAAAGTCAA TCCTGAAAAT ATGCTTTATT
To whom correspondence should be addressed
C3 which is also encoded by C. botulinum C and D phages (4), in particular the C1 neurotoxin lacks a leader sequence which was found in exoenzyme C3.
ACKNOWLEDGEMENTS We are very indebted to P.Cossart and J.Mengaud for providing us with synthetic oligonucleotides. We thank F.Poysky and L.Jordan for help in preparing clostridial phage DNA. This work was supported by grant Nie 175/5-2 from the Deutsche Forschungsgemeinschafts to H.N. (H.K. is a recipient of a fellowship from the Alexander von Humboldt Foundation) and by USAMRIID contract 84-PP-4861-(A-1) to M.E. and by NIH grant A116928 to DMG.
REFERENCES 1. Eklund,M.W., Poysky,F.T., Reed,S.M. and Smith,C.A. (1971) Science 172, 480-482. 2. Eklund,M.W. and Poysky,F.T. (1989) In: Botulinum Neurotoxins and Tetanus Toxin (Simpson,L.L. ed.), pp. 25-51, Academic Press, San Diego. 3. Tsuzuki,K., Yokosawa,N., Syuto,B., Ohishij., Fujii,N., Kimura,K. and Oguma,K. (1988) Infect. Immun. 56, 898-902. 4. Popoff,M.R., Boquet,P., Gill,D.M. and Eklund,M.W. (1990) Nucl. Acids Res. 18, 1291.
1501 TATTTAC74A ATTTT4TCAT AAA6CAATAG ATGGTAGATC ATTATATAAT AAAACATTA6 1561 ATT6TAGA66 6CTTTTA7TT AAA4ATACTG ACTTACCCTT TATAGGT6AT ATt676TAT6 1621 TTAAAACT6A TATATTTrtA AGAA446AT4 TTAAT76GA AACTG67 Tt ATATACTATC 1681 C7647AAT6T TTCA6TAGAT CAAGTTATTC TCAGTAA TA64 TCCTCA6AA CAT67 ACAAC 1741 TAGATTTATT ATACCCT46T ATtGACAGT6 AGAGTGAAAT ATTACCA66 6A6AAATCAA6 1661 TCTTTTAT6A TAATA4476 T C44AAT6TT6 ATTATTTGAA TTCTTATTAT TACCTAG76T 1661 CTCA4AAACT A4676ATAAT 6TTGA467TT TTACTTTTAC 6466TCAATT S66CTT 1921 T66ATAATA6 T6CAAAAGTA TATACTTACT TTCCTACACT AGCtAATAAA 6TAAAT46C6 1981 6T7T4CAA46 T76TTTATTT TTAAT7T476 CAAAT6AT6T A6TTGA46AT TTtACTACAA 2041 ATATtCTAA6 AAAA46TACA TTA6ATAAAA TATCAG46TT ATCAGCTATT ATTCCCT4TA 7 646CATTT6 2101 TA66ACCC6C ATTAAATATA AGTAATTCTG TAAGAAGA76 AAATTTTACT 2161 CAGTTACT66 TOTAACTATT TTATT776 CATTTCCT6A ATTTACAATA CCTGCACTT6 2221 6T6CATTTGT 6ATTTATA7T AA66TTCAAG AAA6 6ACG7 GATTATTAAA ACTATA6ATA 2261 ATt6TTTA6A ACA4467ATT Aa646ATGGA AAGATTCATA TGAATG7TG ATGGGAACGT 2341 G6TTATCCA6 6ATTATTACT CA4TTT6ATAaTATAaGTTA TCAAATGTAT 6ATTCTtTAA 2401 ATTATCA66C A66TBCAATC AAA6CTAAAA TAGATTTAGA ATATAaAAAA TATTCA66AA 2461 6T6AT4A4 AAATATAAAA A6TCAAGTTG AA6ATTTAAA 644TAGTTTA 6aTGTAAAAA 2521 TTtCGAAC AAT4 47TAAT ATAaATAAAT TTATACGAGA AT6TTCC6TA ACATATTTAT 2581 TTAA7AATAT 6TTACCTAA4 6TAATTGAT6 AATTA4AT6A 6TTTBATC6A AATACTAA46 2641 CAAA6ATTAAT TAATCTTATA 6ATATCATA ATATTATTCT A6TT6GTGA6 6TA6ATAAAT 2701 T7AAA6CAAA A6TAAATAAT AGCTTTCAAA ATACAATACC CTtTAATATT TTTTCATATA 4 2761 CTAATAATTC TTTATTA4AA AT466 TATTT CAATAATATT AATGATTCAA ATATAATTA 2621 A4ATTTT646 CCTACAAAAC A4AAAaAATA CTTTAGTG6A TACATCA666 TAT74A7 CAG 2861 Aa6TG7TGA AG644667AT GTTCAGCTTA ATCCAATATT TCCATTTA6C TTTAAATTAG 2941 6TAGTTCAGG 66 46TAGA G6TAAAGTTA TAGTAACCCA GAATGAAA4T ATT6TATATA
3001 ATTCTAT6TA T6A446TTTT A6CATTATT 7 3061 ATTT4CCT7 ATATACTATA ATTGATA6TG 3121 TTA4A767AA T7T7T77 6TA TTTACTTTAA 3181 ATmA6TTA T7ATATATCA AATAATGCTC 3241 TTACTAACAA TATBAT6666 AATAT44764 3301 TAA7A467AA A44ACTAACT 6644TTAATT 3361 AAATTCCA6A TAC766TTTG ATTACTTCAG 3421 ATTTTTATAT ATTT4CT44A 644TTA6AT6 3481 T7CAATATAC T746TTT6TA AAA6ATTATT 3541 ATTATAT66T TAATATA6AT TATTTAAATA 3601 TTTTT4AT4C AC476444T 4ATAATGACT 3661 6 7CA6A4 AAAT4CAAAT GATACTA647 3721 T77 7AATTAA TAACA744 8 TATAATTTGT 3781 ATCATA6TAC T644AGATATA TAT7CTAT8G 3841 ATAATATTAT ATTTCAAATA CAACC7ATG4 3901 TTAAATCAAA TTTTAAT7GA GAAAATATTT 3961 TTA647CTTG A6GTGATT6G TATA6ACACA 4021 ATTAT4CTTC ATTATTA476 TCAACATCAA 4091 AAATAA64AT TAATAATATA AATTATGTTA 4141 TGATTA6TCG AACATTAATA AAATAACTAA 4201 ATAT7TATTA TCAATTACAG TA7TAAAATA 4261 TAAAAA764 6 6ATGTTT74T TTTTA4ATT 4321 TA4A4AATTA AATTAAATT4 TTTT77AAA7 6 4381 CTTATAAAT7 T76644666G ATAAAGC67A 4441 TaTTAAaGGA 6648TTAATA TGGAACACG6
TTTGGATtAG 4ATA4ATAAA TG74GTAAGTA TTAaaAAT4 A CTCA66TT6G A6TATA66TA 4 AACAAAAT6 A AGATAGTG4 CAAAGTATAA CTGGATACAA TaAAT464 TT TTT74AACT6 TTTATATAAA TG6AAAATTA ATA6ATACTA TTA7767CAC TATAACATTT 8667AAATAa ATTCT6ATAA CATCAATAT6 TG6ATAA4A6 GTAAA6ATAT TAATATATTA TTTAATAGCT G6G6464T7A TTTAAGATAT AATAA46aAT GATATAT6TA TGC7 7TCA CGAC4AATTG TCAAT7467 G ATAT4AAATT ATAATAA4A4 TACGA6GAG6 A6ATATTTTA TATTTTGATA TTATGAAGAA T7AAACTAT6 TATGCAGATA 6TTTAA8AGA A6AA76474 6ATATAAATG ATAATACTTA TTATTAC7CA TCTCAAATAT CTGGAATATG TTCAATA66T ACTTATCGTT ATTATTT6GT 6CCTACTGTG AAGCAAGG4A CTCATTG776 7TTTTGTACCT TAA67 TAAT AaTaTTTTaA TATTA467TT AAATATGTTT ATAAATTATA TTTA4AACTG TTAAAATA6 T CT7ATTGCTC TGTCTATAAT CTTCCA6AGT T7TT7466T7
AAGA6ATAC GTTAGCGCAC
'T7TT766CT T74G44A TA TGGTAGTATA TAA7TCTT0G T4CAAAATTA AATTAGAA6 A TC
4924 Nucleic Acids Research, Vol. 18, No. 16
Nucleotide sequence of Clostridium botulinum C1 neurotoxin Daniel Hauser, Melvin W.Eklund2, Hisao Kurazono3, Thomas Binz3, Heiner Niemann 3, D.Michael Gill4, Patrice Boquet1 and Michel R.Popoff* Unite des Anaerobies and 1Unit' des Antigenes Bacteriens, Institut Pasteur, 28 rue du Dr. Roux-75724 Paris cedex 15, France, 2Northwest and Alaska Fisheries Center, 2725 Montlake Boulevard East, Seattle, WA 98112, USA, 3Institut fur Mikrobiologie an der Bundesforschungssanstalt fOr Viruskrankheiten der Tiere, Postfach 1149, 7400 TObingen, FRG and 4Department of Molecular Biology and Microbiology, Tufts University Schools of Medicine, Boston, MA 02111, USA EMBL accession no. X53751
Submitted July 13, 1990
Clostridiu,n botulinum neurotoxins consist of seven serologically distinct types: A, B, C1, D, E, F, and G. All of these are synthesized as single-chain polypeptides of about 150 kDa which are generally nicked after secretion to yield N-terminal light ( = 50 kDa) and C-terminal heavy (= 100 kDa) chains connected by a single disulfide bond. C. botulinum C I neurotoxin is encoded by a bacteriophage (1, 2). C. botulinum Cl neurotoxin DNA sequence shown below was derived from cloned C phage DNA fragments the sizes of which were less than 2.0 kb. The initial clone containing a Ci.neurotoxin coding DNA was identified using a mixed 29-mer probe [5 '-GA(T/C)CCIGTIGA(T/C)AA(T/C)AA(A/G)AA(T/C)ATICTITA-3'] which corresponds to Asp'2 to Tyr21 of the partial amino acid sequence published previously (3). Potential biohazards associated with the experiments described had been approved by the French National Control Committee. The complete C I DNA sequence shows a single open reading frame which begins with an ATG at position 207 and encodes for 1291 amino acids (MR 148,698 Da). Before the ATG a ShineDalgarno sequence can be localized from position 191 to 196. Cys437 and Cys453 are probably involved in a disulfide bridge linking the light and heavy chains. A candidate transcription terminator (AG= -6.2 kCal.mol-1) is underlined. C. botulinum Cl neurotoxin exhibits no similarity with exoenzyme I GAATCAATAG TAAGTCAAAG ATAATGCAAA 61 CT67TTGTAT AACATTTTCA TATAAT6ATA 121 TTATAATTGG ATG6TAT6TA ATGACAATAA 181 TTAGAAAGTT AGGAGATGTT AGTATTATGC 241 ATCCTGTTGA TAATAAAAAT ATTTTA7TATT 301 AGCCTGAAAA AGCCTTTCGC ATTACAGGAA 361 GAAATTCTAA TCCAAATTTA AATAAACCTC 421 ATGATCCTAA TTATTTGT ACTGATTCTG
481 A4TTATTTAA AA4AATTAAT 541 CAGATATACC CTTTCCTGGG
7TATATGGGT AATACCT6AT AGATTTTC6 A
CTCGAGTTAC AAGCCCTAAA AGTGGTTATT ACAAAGATCC ATTTTTAAAA GAAATTATAA TCTAGAGA4A TAG6AGAAGA ATTAATATAT AGACTTTCGA AATAACAATA CTCCAATT'A T6CTTTTGAT TTTGATGTA6 AAAACTA6AC AAGGTAACAA CT68TTAA4 ACT6GTA6CA ACTGGACCTA G4GAAAACAT TATAGATCCA GAAACTTCTA ACTTTTGC6G 7ACAAGA4GG ATTTGT76CT TTATCAATAA ATGCTAACAT ATAGTAATGC 7ACTAAT6AT GTA46G6 TTTTCATG66 ATCCAATACT AaTTTTAAT6 CATGAACTTA TATG847A8G CTATACCAAA TGATCAAACA ATTTCATCT6 TCTCAATATA AT7TGA6ATT AG6ATATGCA 6AAATATATG
601 ATTTTAACA66 TTTGATGTT 661 TAAATCCTAG T6TTATAATA 721 C8TTTAAATT AACTAACAAT 781 TTTCAATATC ACCTAGATTT 841 6TAGATTT7C TAAGTCTGAA 901 ATCATGCAAT 6CATAATTTA 961 TAACTAGTAA TATTTTTTAT 1021 CATTT7766 TCCAACTATA G4CCTTATTC 1081 A4 766ATTG6A TATTATAGA TCTATAGCTA 1141 CTTCAAGCTT T7ATAAAAT4AT7A466844AT 1201 TC7TA6TA6A ATCTTCAGGT TTACAG 1261 ATAA4CTTAC ACAAATATTT ACA766TTTA 1321 B6AAaATA6 A TCTTTCAAAT 6TATATACTC 1361 TtTAT7A6AT ACAAAAT76 TTtAATATAC 1441 6TCAA4AT4 T ATCTC6AAAT CCA4CATTAA *
AAGATTACAG TTAATACTGA TTATTTA6AC AATATT7TCT CTATC4CTTA GAGATGGAGA CAAG6TGCTA AAGGTGCACA TTTGTGGATA CAATAACAAT TAACAACTTT AATTATTCAG TAGATACTCA TTTAAATACA CTAGCTAATG
CTAAA76TGC AA6BAAATAT TTTGA7664
AAAGACTTAA TAGTATAACT ACTGCAAATC ATA6 7AGAA ACTTATTA6A AA6TATA64T
TAAATCGTAA TAA6TTTGTT 946TTAT6TA ACTACGCTAA AATATATAAT 6TACAA4ATA C6GTT6C6GC 8AATATATTA 64CGATAAT6
CTAAAA6TAA TTTAAAT7TA CTATTTAT76
GAAAAGTCAA TCCTGAAAAT ATGCTTTATT
To whom correspondence should be addressed
C3 which is also encoded by C. botulinum C and D phages (4), in particular the C1 neurotoxin lacks a leader sequence which was found in exoenzyme C3.
ACKNOWLEDGEMENTS We are very indebted to P.Cossart and J.Mengaud for providing us with synthetic oligonucleotides. We thank F.Poysky and L.Jordan for help in preparing clostridial phage DNA. This work was supported by grant Nie 175/5-2 from the Deutsche Forschungsgemeinschafts to H.N. (H.K. is a recipient of a fellowship from the Alexander von Humboldt Foundation) and by USAMRIID contract 84-PP-4861-(A-1) to M.E. and by NIH grant A116928 to DMG.
REFERENCES 1. Eklund,M.W., Poysky,F.T., Reed,S.M. and Smith,C.A. (1971) Science 172, 480-482. 2. Eklund,M.W. and Poysky,F.T. (1989) In: Botulinum Neurotoxins and Tetanus Toxin (Simpson,L.L. ed.), pp. 25-51, Academic Press, San Diego. 3. Tsuzuki,K., Yokosawa,N., Syuto,B., Ohishij., Fujii,N., Kimura,K. and Oguma,K. (1988) Infect. Immun. 56, 898-902. 4. Popoff,M.R., Boquet,P., Gill,D.M. and Eklund,M.W. (1990) Nucl. Acids Res. 18, 1291.
1501 TATTTAC74A ATTTT4TCAT AAA6CAATAG ATGGTAGATC ATTATATAAT AAAACATTA6 1561 ATT6TAGA66 6CTTTTA7TT AAA4ATACTG ACTTACCCTT TATAGGT6AT ATt676TAT6 1621 TTAAAACT6A TATATTTrtA AGAA446AT4 TTAAT76GA AACTG67 Tt ATATACTATC 1681 C7647AAT6T TTCA6TAGAT CAAGTTATTC TCAGTAA TA64 TCCTCA6AA CAT67 ACAAC 1741 TAGATTTATT ATACCCT46T ATtGACAGT6 AGAGTGAAAT ATTACCA66 6A6AAATCAA6 1661 TCTTTTAT6A TAATA4476 T C44AAT6TT6 ATTATTTGAA TTCTTATTAT TACCTAG76T 1661 CTCA4AAACT A4676ATAAT 6TTGA467TT TTACTTTTAC 6466TCAATT S66CTT 1921 T66ATAATA6 T6CAAAAGTA TATACTTACT TTCCTACACT AGCtAATAAA 6TAAAT46C6 1981 6T7T4CAA46 T76TTTATTT TTAAT7T476 CAAAT6AT6T A6TTGA46AT TTtACTACAA 2041 ATATtCTAA6 AAAA46TACA TTA6ATAAAA TATCAG46TT ATCAGCTATT ATTCCCT4TA 7 646CATTT6 2101 TA66ACCC6C ATTAAATATA AGTAATTCTG TAAGAAGA76 AAATTTTACT 2161 CAGTTACT66 TOTAACTATT TTATT776 CATTTCCT6A ATTTACAATA CCTGCACTT6 2221 6T6CATTTGT 6ATTTATA7T AA66TTCAAG AAA6 6ACG7 GATTATTAAA ACTATA6ATA 2261 ATt6TTTA6A ACA4467ATT Aa646ATGGA AAGATTCATA TGAATG7TG ATGGGAACGT 2341 G6TTATCCA6 6ATTATTACT CA4TTT6ATAaTATAaGTTA TCAAATGTAT 6ATTCTtTAA 2401 ATTATCA66C A66TBCAATC AAA6CTAAAA TAGATTTAGA ATATAaAAAA TATTCA66AA 2461 6T6AT4A4 AAATATAAAA A6TCAAGTTG AA6ATTTAAA 644TAGTTTA 6aTGTAAAAA 2521 TTtCGAAC AAT4 47TAAT ATAaATAAAT TTATACGAGA AT6TTCC6TA ACATATTTAT 2581 TTAA7AATAT 6TTACCTAA4 6TAATTGAT6 AATTA4AT6A 6TTTBATC6A AATACTAA46 2641 CAAA6ATTAAT TAATCTTATA 6ATATCATA ATATTATTCT A6TT6GTGA6 6TA6ATAAAT 2701 T7AAA6CAAA A6TAAATAAT AGCTTTCAAA ATACAATACC CTtTAATATT TTTTCATATA 4 2761 CTAATAATTC TTTATTA4AA AT466 TATTT CAATAATATT AATGATTCAA ATATAATTA 2621 A4ATTTT646 CCTACAAAAC A4AAAaAATA CTTTAGTG6A TACATCA666 TAT74A7 CAG 2861 Aa6TG7TGA AG644667AT GTTCAGCTTA ATCCAATATT TCCATTTA6C TTTAAATTAG 2941 6TAGTTCAGG 66 46TAGA G6TAAAGTTA TAGTAACCCA GAATGAAA4T ATT6TATATA
3001 ATTCTAT6TA T6A446TTTT A6CATTATT 7 3061 ATTT4CCT7 ATATACTATA ATTGATA6TG 3121 TTA4A767AA T7T7T77 6TA TTTACTTTAA 3181 ATmA6TTA T7ATATATCA AATAATGCTC 3241 TTACTAACAA TATBAT6666 AATAT44764 3301 TAA7A467AA A44ACTAACT 6644TTAATT 3361 AAATTCCA6A TAC766TTTG ATTACTTCAG 3421 ATTTTTATAT ATTT4CT44A 644TTA6AT6 3481 T7CAATATAC T746TTT6TA AAA6ATTATT 3541 ATTATAT66T TAATATA6AT TATTTAAATA 3601 TTTTT4AT4C AC476444T 4ATAATGACT 3661 6 7CA6A4 AAAT4CAAAT GATACTA647 3721 T77 7AATTAA TAACA744 8 TATAATTTGT 3781 ATCATA6TAC T644AGATATA TAT7CTAT8G 3841 ATAATATTAT ATTTCAAATA CAACC7ATG4 3901 TTAAATCAAA TTTTAAT7GA GAAAATATTT 3961 TTA647CTTG A6GTGATT6G TATA6ACACA 4021 ATTAT4CTTC ATTATTA476 TCAACATCAA 4091 AAATAA64AT TAATAATATA AATTATGTTA 4141 TGATTA6TCG AACATTAATA AAATAACTAA 4201 ATAT7TATTA TCAATTACAG TA7TAAAATA 4261 TAAAAA764 6 6ATGTTT74T TTTTA4ATT 4321 TA4A4AATTA AATTAAATT4 TTTT77AAA7 6 4381 CTTATAAAT7 T76644666G ATAAAGC67A 4441 TaTTAAaGGA 6648TTAATA TGGAACACG6
TTTGGATtAG 4ATA4ATAAA TG74GTAAGTA TTAaaAAT4 A CTCA66TT6G A6TATA66TA 4 AACAAAAT6 A AGATAGTG4 CAAAGTATAA CTGGATACAA TaAAT464 TT TTT74AACT6 TTTATATAAA TG6AAAATTA ATA6ATACTA TTA7767CAC TATAACATTT 8667AAATAa ATTCT6ATAA CATCAATAT6 TG6ATAA4A6 GTAAA6ATAT TAATATATTA TTTAATAGCT G6G6464T7A TTTAAGATAT AATAA46aAT GATATAT6TA TGC7 7TCA CGAC4AATTG TCAAT7467 G ATAT4AAATT ATAATAA4A4 TACGA6GAG6 A6ATATTTTA TATTTTGATA TTATGAAGAA T7AAACTAT6 TATGCAGATA 6TTTAA8AGA A6AA76474 6ATATAAATG ATAATACTTA TTATTAC7CA TCTCAAATAT CTGGAATATG TTCAATA66T ACTTATCGTT ATTATTT6GT 6CCTACTGTG AAGCAAGG4A CTCATTG776 7TTTTGTACCT TAA67 TAAT AaTaTTTTaA TATTA467TT AAATATGTTT ATAAATTATA TTTA4AACTG TTAAAATA6 T CT7ATTGCTC TGTCTATAAT CTTCCA6AGT T7TT7466T7
AAGA6ATAC GTTAGCGCAC
'T7TT766CT T74G44A TA TGGTAGTATA TAA7TCTT0G T4CAAAATTA AATTAGAA6 A TC
