diff --git a/Snakefile.genome b/Snakefile.genome index ea7ecdf..d32222b 100644 --- a/Snakefile.genome +++ b/Snakefile.genome @@ -74,6 +74,7 @@ rule align: --reference-sequence {input.reference} \ --output {output.alignment} \ --remove-reference \ + --fill-gaps \ --nthreads {threads} """ @@ -188,6 +189,7 @@ rule ancestral: input: tree = "results/{build_name}/genome/tree.nwk", alignment = "results/{build_name}/genome/aligned.fasta", + root_sequence = "config/h5_cattle_genome_root.gb", output: node_data = "results/{build_name}/genome/nt-muts.json" params: @@ -198,7 +200,8 @@ rule ancestral: --tree {input.tree} \ --alignment {input.alignment} \ --output-node-data {output.node_data} \ - --inference {params.inference}\ + --inference {params.inference} \ + --root-sequence {input.root_sequence} \ --keep-ambiguous """ @@ -207,7 +210,7 @@ rule translate: input: tree = "results/{build_name}/genome/tree.nwk", node_data = "results/{build_name}/genome/nt-muts.json", - reference = "results/{build_name}/genome/reference.gb", + reference = "config/h5_cattle_genome_root.gb", output: node_data = "results/{build_name}/genome/aa-muts.json" shell: @@ -219,6 +222,7 @@ rule translate: --output {output.node_data} """ + rule traits: message: "Inferring ancestral traits for {params.columns!s}" input: diff --git a/config/auspice_config_h5n1-cattle-outbreak.json b/config/auspice_config_h5n1-cattle-outbreak.json index f4bf5de..164494a 100755 --- a/config/auspice_config_h5n1-cattle-outbreak.json +++ b/config/auspice_config_h5n1-cattle-outbreak.json @@ -14,6 +14,31 @@ "name": "GISAID" } ], + "extensions": { + "nextclade": { + "pathogen": { + "schemaVersion":"3.0.0", + "defaultCds": "HA", + "cdsOrderPreference":[ + "PB2", + "PB1", + "PA", + "HA", + "NP", + "NA", + "M1", + "M2", + "NS1", + "NS2" + ], + "attributes": { + "name": "H5N1 dairy cattle outbreak", + "reference name": "concatenated ancestral sequences", + "reference accession": "none" + } + } + } + }, "colorings": [ { "key": "gt", diff --git a/config/h5_cattle_genome_root.gb b/config/h5_cattle_genome_root.gb new file mode 100644 index 0000000..ba0cfc5 --- /dev/null +++ b/config/h5_cattle_genome_root.gb @@ -0,0 +1,383 @@ +LOCUS inferred_root_sequence 13590 bp RNA linear VRL 2024 +DEFINITION Combined references for 8 segments of the inferred ancestral sequence of the 2024 US dairy cattle outbreak. +ACCESSION XXXXXXXX +VERSION XXXXXXXX.1 +KEYWORDS . +SOURCE Influenza A virus (coordinates from A/goose/Guangdong/1/1996(H5N1) AF144300) + ORGANISM Influenza A virus (coordinates from A/goose/Guangdong/1/1996(H5N1) AF144300) + Viruses; ssRNA viruses; ssRNA negative-strand viruses; + Orthomyxoviridae; Influenzavirus A. +FEATURES Location/Qualifiers + source 1..13590 + /organism="Influenza A virus + (A/goose/Guangdong/1/1996(H5N1))" + /mol_type="genomic RNA" + /strain="A/Goose/Guangdong/1/96(H5N1)" + /db_xref="taxon:93838" + CDS 28..2307 + /gene="PB2" + /codon_start=1 + /product="polymerase" + /protein_id="AAD51922.1" + /translation="MERIKELRDLMSQSRTREILTKTTVDHMAIIKKYTSGRQEKNPAL + RMKWMMAMKYPIAADKRIMEMIPERNEQGQTLWSKTNDAGSDRVMVSPLAVTWWNRNGP + TTSTIHYPKVYKTYFEKVERLKHGTFGPVHFRNQIKIRRRVDINPGHADLSAKEAQDVI + MEVVFPNEVGARILTSESQLTITKEKKEELQDCKIAPLMVAYMLERELVRKTRFLPVAG + GTSSVYIEVLHLTQGTCWEQMYTPGGEVRNDDVDQSLIIAARNIVRRATVSADPLASLL + EMCHSTQIGGIRMVDILRQNPTEEQAVDICKAAMGLRISSSFSFGGFTFKRTSGSSVKR + EEEVLTGNLQTLKIRVHEGYEEFTMVGRRATAILRKATRRLIQLIVSGRDEQSIAEAII + VAMVFSQEDCMIKAVRGDLNFVNRANQRLNPMHQLLRHFQKNAKVLFQNWGIEPIDNVM + GMIGILPDMTPSTEMSLRGIRVSKMGVDEYSSTERVIVSIDRFLRVRDQRGNVLLSPEE + VSETQGTEKLTITYSSSMMWEINGPESVLVNTYQWIIRNWETVKIQWSQDPTMLYNKME + FEPFQSLVPKAARGQYSGFVRTLFQQMRDVLGTFDTVQIIKLLPFAAAPPEQSRMQFSS + LTVNVRGSGMRILIRGNSPVFNYNKATKRLTVLGKDAGALAEDPDEGTAGVESAVLRGF + LILGKEDKRYGPALSINELSNLAKGEKANVLIGQGDVVLVMKRKRDSSILTDSQTATKR + IRMAIN" + CDS 2366..4639 + /gene="PB1" + /codon_start=1 + /product="polymerase" + /protein_id="AAD51923.1" + /translation="MDVNPTLLFLKVPAQNAISTTFPYTGDPPYSHGTGTGYTMDTVNR + THQYSEKGKWTTNSETGAPQLNPIDGPLPDDNEPSGYAQTDCVLEAMAFLEESHPGIFE + NSCLETMEVVQQTRVDKLTQGRQTYDWTLNRNQPAATALANTIEVFRSNGLTANESGRL + IDFLKDVVESMDKEEIEITTHFQRKRRVRDNMTKKMVTQRTIGKKKQRLNKRSYLIRAL + TLNTMTKDAERGKLKRRAIATPGMQIRGFVYFVETLARSICEKLEQSGLPVGGNEKKAK + LANVVRKMMTNSQDTELSFTITGDNTKWNENQNPRMFLAMITYITRNQPEWFRNVLSIA + PIMFSNKMARLGKGYMFESKSMKLRTQIPAEMLASIDLKYFNESTRKKIEKIRPLLIDG + TASLSPGMMMGMFNMLSTVLGVSILNLGQKKYTKTTYWWDGLQSSDDFALIVNAPNHEG + IQAGVDRFYRTCKLVGINMSKKKSYINRTGTFEFTSFFYRYGFVANFSMELPSFGVSGI + NESADMSIGVTVIKNNMINNDLGPATAQMALQLFIKDYRYTYRCHRGDTQIQTRRSFEL + KKLWEQTRSKPGLLVSDGGPNLYNIRNLHIPEVCLKWELMDEDYQGRLCNPLNPFVSHK + EIESVNNAVVMPAHGPAKSMEYDAVATTHSWIPKRNRSILNTSQRGILEDEQMYQKCCN + LFEKFFPSSSYRRPVGISSMVEAMVSRARIDARIDFESGRIKKEEFAEIMKICSTIEEL + RRQK" + CDS 4707..6857 + /gene="PA" + /codon_start=1 + /product="polymerase" + /protein_id="AAD51924.1" + /translation="MEDFVRQCFNPMIVELAEKAMKEYGEDPKIETNKFAAICTHLEVC + FMYSDFHFIDERGESMIVESGDPNALLKHRFEIIEGRDRAMAWTVVNSICNTTGVEKPK + FLPDLYDYRENRFIEIGVTRREVHIYYLEKANKIKSEKTHIHIFSFTGEEMATKADYTL + DEESRARIKTRLFTIRQEMASRGLWDSFRQSERGEETIEERFEITGTMRRLADQSLPPN + FSSLENFRAYVDGFEPNGCIEGKLSQMSKEVNARIEPFLKTTPRPLRLPDGPPCPQRSK + FLLMDALKLSIEDPSHEGEGIPLYDAIKCMKTFFGWKEPNIVKPHEKGINPNYLLAWKQ + VLAELQDIENEEKIPKTKNMKKTSQLKWALGENMAPEKVDFEDCKDVSDLRQYDSDEPE + SRSLASWIQSEFNKACELTDSSWIELDEIGEDVAPIEHIASVRRNYFTAEVSHCRATEY + IMKGVYINTALLNASCAAMDDFQLIPMISKCRTKEGRRKTNLYGFIIKGRSHLRNDTDV + VNFVSMEFSLTDPRLEPHKWEKYCVLEIGDMLLRTAIGQVLRPMFLYVRTNGTSKIKMK + WGMEMRRCLLQSLQQIESMIEAESSVKEKDMSKEFFENKSETWPIGESPKGVEEGSIGK + VCRTLLAKSVFNSLYASPQLEGFSAESRKLLLIVQALRDNLEPGTFDLGGLYEAIEECL + INDPWVLLNASWFNSFLTHALK" + CDS 6937..8643 + /gene="HA" + /codon_start=1 + /product="hemagglutinin" + /protein_id="AAD51927.1" + /translation="MENIVLLLAIVSLVKSDQICIGYHANNSTEQVDTIMEKNVTVTHA + QDILEKTHNGKLCDLNGVKPLILKDCSVAGWLLGNPMCDEFIRVPEWSYIVERANPAND + LCYPGSLNDYEELKHMLSRINHFEKIQIIPKSSWPNHETSLGVSAACPYQGAPSFFRNV + VWLIKKNDAYPTIKISYNNTNREDLLILWGIHHSNNAEEQTNLYKNPITYISVGTSTLN + QRLAPKIATRSQVNGQRGRMDFFWTILKPDDAIHFESNGNFIAPEYAYKIVKKGDSTIM + KSGVEYGHCNTKCQTPVGAINSSMPFHNIHPLTIGECPKYVKSNKLVLATGLRNSPLRE + KRRKXRGLFGAIAGFIEGGWQGMVDGWYGYHHSNEQGSGYAADKESTQKAIDGVTNKVN + SIIDKMNTQFEAVGREFNNLERRIENLNKKMEDGFLDVWTYNAELLVLMENERTLDFHD + SNVKNLYDKVRLQLRDNAKELGNGCFEFYHKCDNECMESVRNGTYDYPQYSEEARLKRE + EISGVKLESVGTYQILSIYSTAASSLALAIMMAGLSLWMCSNGSLQCRICI" + CDS 8721..10217 + /gene="NP" + /codon_start=1 + /product="nucleocapsid protein" + /protein_id="AAD51925.1" + /translation="MASQGTKRSYEQMETGGERQNATEIRASVGRMVGGIGRFYIQMCT + ELKLSDHEGRLIQNSITIERMVLSAFDERRNKYLEEHPSAGKDPKKTGGPIYRRRDGKW + MRELILYDKEEIRRIWRQANNGEDATAGLTHLMIWHSNLNDATYQRTRALVRTGMDPRM + CSLMQGSTLPRRSGAAGAAVKGVGTMVMELIRMIKRGINDRNFWRGENGRRTRIAYERM + CNILKGKFQTAAQRAMMDQVRESRNPGNAEIEDLIFLARSALILRGSVAHKSCLPACVY + GLAVASGYDFEREGYSLVGIDPFRLLQNSQVFSLIRPNENPAHKSQLVWMACHSAAFED + LRVSSFIRGTRVVPRGQLSTRGVQIASNENMETMDSSTLELRSRYWAIRTRSGGNTNQQ + RASAGQISVQPTFSVQRNLPFERATIMAAFTGNTEGRTSDMRTEIIRMMENARPEDVSF + QGRGVFELSDEKATNPIVPSFDMNNEGSYFFGDNAEEYDN" + CDS 10261..11670 + /gene="NA" + /codon_start=1 + /product="neuraminidase" + /protein_id="AAD51926.1" + /translation="MNPNQKITTIGSICMVIGIVSLMLQIGNIISIWVSHSIQTGNQYQ + PEPCNQSIITYENNTWVNQTYINISNTNFLAEQAVTSVTLAGNSSLCPISGWAIYSKDN + GIRIGSKGDVFVIREPFISCSHLECRTFFLTQGALLNDKHSNGTVKDRSPYRTLMSCPV + GEAPSPYNSRFESVAWSASACHDGISWLTIGISGPDNGAVAVLKYNGIITDTIKSWRNN + ILRTQESECACVNGSCFTVMTDGPSNGQASYKIFKIEKGKVVKSVEMNAPNYHYEECSC + YPDAGDIMCVCRDNWHGSNRPWVSFNQNLEYQIGYICSGIFGDNPRPNDGTGSCSPMPS + NGAYGVKGFSFKYGNGVWIGRTKSTSSRSGFEMIWDPNGWTETDSSFSVKQDIVEITDW + SGYSGSFVQHPELTGLDCMRPCFWVELIRGRPKENTIWTSGSSISFCGVNSDTVGWSWP + DGAELPFTIDK" + CDS join(11724..11749,12438..12705) + /gene="M2" + /note="M2" + /codon_start=1 + /product="matrix protein 2" + /protein_id="AAD51929.1" + /translation="MSLLTEVETPTKNGWECNCSDSSDPLVIAASIIGILHLILWILDR + LFFKCVYRRLKYGLKGGPSTEGVPESMREEYRQEQQSAVDVDNGHFVNIELE" + CDS 11724..12482 + /gene="M1" + /note="M1" + /codon_start=1 + /product="matrix protein 1" + /protein_id="AAD51928.1" + /translation="MSLLTEVETYVLSIVPSGPLKAEIAQRLEDVFAGKNTDLEALMEW + LKTRPILSPLTKGILGFVFTLTVPSERGLQRRRFVQSALSGTGDPNNMDRAVKLYRKLK + REITFHGAKEVALSYSTGALASCMGLIYNRMGTVTAEVALGLVCATCEQIADSQHRSHR + QIATTTNPLIRHENRMVLASTTAKAMEQMAGSSEQAVEAMEVASQARQMVQAMRTIGTH + PSSSTGLRDDLLENLQAYQKRMGVQLQRFK" + CDS join(12740..12769,13242..13577) + /gene="NS2" + /note="NS2" + /codon_start=1 + /product="nonstructural protein 2" + /protein_id="AAD51931.1" + /translation="MDSNTVLSFQDILMRMSKMQLGSSSEDLNGMITQFESLKLYRDSL + GEAVMRMGDLHSLQSRNGKWRGQLSQKFEEIRWLIEEVRHRLKITENSFEQITFMQALQ + LLLEVEQEIRTFSFQLI" + CDS 12740..13432 + /gene="NS1" + /note="NS1" + /codon_start=1 + /product="nonstructural protein 1" + /protein_id="AAD51930.1" + /translation="MDSNTVLSFQVDCFLWHVRKRFADQELGDAPFLDRLRRDQKSLRG + RGSTLGLDIETATRAGKQIVERILEEESDEALKMTIASVPAPRYLTDMTLEEMSRDWFM + LMPKQKVAGSLSIRMDQAIMDKNIILKANFSVIFNRLETLILLRAFTEEGAIVGEISPL + PSLPGHTDEDVKNAIGVLIGGLEWNDNTVRVSETLQRFAWRSSNEDGRPPLPPKQKRKM + ERTIESEV" +ORIGIN + 1 agcaaaagca ggtcaaatat attcaatatg gagagaataa aagaactgag agatctaatg + 61 tcacagtctc gcactcgcga gatactaacc aaaaccactg ttgaccacat ggccataatc + 121 aagaagtaca catcaggaag acaagaaaag aaccctgcac tcagaatgaa atggatgatg + 181 gcaatgaaat atccaatcgc agcagacaag cgaataatgg aaatgatccc tgaaaggaat + 241 gaacaaggac aaaccctctg gagcaagaca aatgatgccg gatcagatcg agtgatggta + 301 tcacccctgg ctgtgacatg gtggaatagg aatggaccaa caacaagtac aattcactat + 361 ccaaaggtat acaaaactta ttttgaaaaa gttgaaaggt tgaaacacgg gacctttggc + 421 cctgtacact tcagaaacca aattaagata agacggaggg tcgacataaa cccgggccat + 481 gctgacctca gcgccaaaga ggcgcaggac gtaatcatgg aagttgtctt tccaaatgaa + 541 gtgggagcga gaatactgac gtcggaatca caattgacaa taacaaagga aaagaaagaa + 601 gaactccagg actgcaaaat cgcccctctg atggttgcat acatgctaga aagagagctg + 661 gtccgcaaga caaggttcct cccagttgct ggtggaacaa gcagtgtcta cattgaggtg + 721 ctgcatttga cccagggaac atgctgggag cagatgtata ctccaggagg agaagtgaga + 781 aacgatgatg tagaccagag cttgatcatt gctgccagga atatagtaag aagagcaaca + 841 gtgtcagcag acccattagc atctctattg gagatgtgcc acagcacaca aattggagga + 901 ataaggatgg tagacattct tcggcaaaat ccaacggagg aacaagccgt ggacatatgc + 961 aaggcagcaa tgggcttgag gattagctca tcgttcagct ttggtggatt cacttttaaa + 1021 agaacaagtg gatcatcagt caaaagggaa gaagaggtgc ttacgggcaa ccttcaaaca + 1081 ttgaaaataa gagtacatga ggggtatgaa gagttcacta tggttggaag aagagcaacg + 1141 gccattctca ggaaagcaac cagaaggctg attcagctaa tagtaagtgg aagggacgaa + 1201 cagtcaattg ctgaagcaat aatcgtggcc atggtattct cacaagagga ctgcatgata + 1261 aaggcagttc gaggtgatct gaactttgtc aacagggcga atcagcggct gaatccaatg + 1321 catcagctct tgagacactt ccaaaagaat gcaaaagtgc ttttccaaaa ttggggaatt + 1381 gagcccattg acaatgtgat gggaatgatc gggatattgc ctgacatgac tccaagtact + 1441 gagatgtcac tgaggggaat aagagtcagt aagatgggag tagatgaata ctccagtaca + 1501 gagcgggtaa tagtaagcat cgaccgattt ttaagagttc gagaccaacg ggggaacgta + 1561 ctactgtcac ccgaagaagt cagcgagaca caaggaacag agaaattgac aatcacttat + 1621 tcgtcatcaa tgatgtggga gatcaatggt cctgagtcgg tgttggtcaa tacttatcag + 1681 tggataatca gaaactggga aacagtaaaa attcaatggt cacaagatcc cacaatgttg + 1741 tataataaga tggagttcga gccattccag tctctggtcc ctaaggcagc caggggtcaa + 1801 tacagtgggt tcgtgaggac actatttcag caaatgcgag atgtgcttgg aacatttgac + 1861 actgttcaga taataaaact tctccccttt gctgctgccc caccggaaca aagtagaatg + 1921 caattctcct ctctgactgt gaatgtgaga ggatcaggaa tgagaatact gataagaggc + 1981 aattctccag tgttcaatta caacaaggcc accaagaggc tcacagttct cgggaaagat + 2041 gcaggtgcat tggccgaaga tccagatgaa ggcacagctg gagtggagtc tgctgtttta + 2101 agaggattcc tcattttggg caaagaagac aagagatatg gcccagcact gagcatcaat + 2161 gagctgagca atttggcaaa gggagagaag gctaatgtgc taattgggca aggagacgtg + 2221 gtgttggtga tgaaacggaa acgggactct agcatactta ctgacagcca gacagcgacc + 2281 aaaagaattc ggatggccat caattagtgt agaattgttt aaaaacgacc ttgtttctac + 2341 tagcaaaagc aggcaaacca tttgaatgga tgtcaatccg accttactct tcttgaaagt + 2401 tccagcgcaa aatgccataa gcaccacatt cccgtatact ggagatcctc catacagcca + 2461 tggaacagga acaggatata ccatggacac agttaacaga acacatcaat attcagaaaa + 2521 agggaaatgg acaacaaact cagaaaccgg ggcacctcaa ctcaatccaa ttgatggacc + 2581 attgcctgat gacaatgagc caagtggata tgcacaaacg gactgcgtcc ttgaagcaat + 2641 ggctttcctt gaagaatccc atccaggaat ctttgaaaac tcgtgtcttg aaacgatgga + 2701 agttgttcaa caaacaagag tggacaagtt gacccaaggc cgtcagactt atgattggac + 2761 attaaacaga aatcagccgg ctgcaactgc attagctaat actatagagg tcttcagatc + 2821 gaacggtctt acagctaatg aatcaggaag gctaatagat ttcctcaagg atgtggtgga + 2881 atcaatggat aaagaggaaa tagaaataac aacgcatttc caaaggaaaa gaagagtgag + 2941 agacaacatg accaagaaaa tggtcacaca acggacgata ggaaagaaga aacaaaggtt + 3001 aaacaaaagg agctatctga taagagcatt gacactgaac acaatgacaa aagacgccga + 3061 aagaggcaaa ttaaagagaa gggcaattgc aacacccgga atgcaaatca gagggtttgt + 3121 gtactttgtt gaaacattag caaggagcat ttgtgagaaa cttgaacaat ctggactccc + 3181 agttggaggc aatgaaaaga aggccaaact agcaaatgtt gtgagaaaga tgatgactaa + 3241 ttcgcaagac acagagctct ctttcacaat cacgggagac aacactaaat ggaatgagaa + 3301 ccagaatcct aggatgtttc tggcaatgat aacatatata acaaggaacc aacctgaatg + 3361 gttcaggaat gtattgagca ttgcacctat aatgttctca aacaaaatgg caagactagg + 3421 gaaaggatac atgttcgaaa gtaagagcat gaagcttcga acacaaatac cggcagaaat + 3481 gctagcgagc attgatctga aatacttcaa tgagtcaaca aggaagaaaa tagagaagat + 3541 aagacctctt ctaatagatg gtacggcctc attaagccct ggaatgatga tgggcatgtt + 3601 caacatgctg agtacagttc tgggagtttc gattctaaat ctagggcaaa agaagtacac + 3661 caaaacaaca tactggtggg atggactaca atcttctgat gactttgctc tcatcgtgaa + 3721 tgctccaaat catgagggaa tacaagcagg agtagacaga ttctatagaa cctgcaagct + 3781 ggtaggaatc aatatgagca aaaagaagtc atacataaac aggacaggaa catttgaatt + 3841 cacaagtttt ttctatcgct atggatttgt agccaatttc agcatggagt tgcccagctt + 3901 tggagtttct gggatcaatg aatctgcaga catgagcatt ggagtaacag tgataaagaa + 3961 caacatgatc aacaatgatc ttggaccagc aacagcccaa atggctctac agctattcat + 4021 caaggattac agatacacat atcgatgtca cagaggagac acacaaattc aaacaaggag + 4081 gtcattcgag ctgaaaaagt tatgggaaca aacccgctca aaaccaggac tgctggtctc + 4141 agatggaggg ccaaatctat acaatatccg aaatctccac attccggaag tctgcttaaa + 4201 atgggagcta atggacgaag actatcaggg aaggctttgt aatcccctga atccgtttgt + 4261 aagccacaaa gaaatagagt ctgtgaacaa tgctgtggtg atgccagctc atggcccagc + 4321 taagagtatg gaatatgatg ctgttgccac cactcactcc tggatcccta agaggaaccg + 4381 ctctattctt aatacaagcc aaaggggaat ccttgaagac gaacagatgt atcaaaagtg + 4441 ctgcaatcta tttgaaaaat tcttccctag cagttcatac aggaggccgg ttggaatttc + 4501 cagcatggtg gaggccatgg tttctagggc ccgaattgat gcacgaattg acttcgaatc + 4561 tggacggatt aagaaggagg agtttgctga gatcatgaag atctgttcca ccattgaaga + 4621 gctcagacgg cagaaatagt gaatttagct tgtccttcat gaaaaaatgc cttgtttcta + 4681 ctagcaaaag caggtactga ttcaaaatgg aagactttgt gcgacaatgc ttcaatccaa + 4741 tgattgtcga gcttgcggaa aaagcaatga aagaatatgg ggaagatccg aaaatcgaga + 4801 caaacaaatt tgccgcaata tgcacacact tagaagtctg tttcatgtat tcggatttcc + 4861 attttattga cgaacgaggc gaatcaatga ttgtagaatc tggcgatcca aatgcattat + 4921 tgaaacaccg atttgagata atcgaaggga gagaccgagc aatggcctgg acagtggtga + 4981 atagtatctg caacaccaca ggggtcgaaa agcccaaatt cctccctgat ttgtatgact + 5041 acagagagaa cagattcatt gaaattggag taacgcgaag ggaagttcac atatactatt + 5101 tggaaaaagc caacaagata aaatcagaga aaacacatat tcacatattc tcattcactg + 5161 gagaggaaat ggccaccaag gcggactaca cccttgatga agagagcaga gcaagaataa + 5221 aaaccagact gttcactata agacaagaaa tggccagtag aggtctatgg gattcctttc + 5281 gtcaatccga gagaggcgaa gagacaattg aagaaagatt tgaaatcaca ggaaccatgc + 5341 gcaggcttgc cgaccaaagt cttccaccga acttctccag ccttgaaaac tttagagcct + 5401 atgtggatgg attcgaaccg aacggctgca ttgagggcaa gctttctcaa atgtcaaaag + 5461 aggtgaacgc cagaattgag ccatttctga agacaacacc acgccctctc agattacctg + 5521 atgggcctcc ctgtcctcag cggtcgaagt tcttgctgat ggatgccctt aagttgagca + 5581 tcgaagaccc tagtcatgag ggggagggca taccgctgta tgatgcaatc aaatgcatga + 5641 agacattttt tggctggaaa gagcccaaca tcgtaaagcc gcatgagaaa ggcataaacc + 5701 ctaattacct cctggcttgg aagcaggtgc tggcagaact tcaagacatt gaaaatgagg + 5761 agaaaattcc aaaaacaaag aacatgaaga aaacaagcca attgaagtgg gcacttggtg + 5821 agaacatggc tccagaaaaa gtggactttg aggactgcaa agatgttagc gatctaagac + 5881 agtacgacag tgacgaacca gagtctagat cactagcaag ctggattcag agtgaattca + 5941 acaaggcatg cgaactgaca gattcgagtt ggattgaact tgatgagata ggggaagacg + 6001 ttgctccaat cgaacacatt gcgagtgtga ggaggaacta tttcacagcg gaggtatccc + 6061 attgcagggc cactgaatac ataatgaagg gagtatacat aaacacagcc ctattgaatg + 6121 catcctgtgc agccatggat gacttccaat tgattccaat gataagtaag tgcagaacta + 6181 aagaaggaag acggaagaca aatctgtatg gattcattat aaaaggaaga tcccatttga + 6241 ggaatgacac cgatgtggta aactttgtga gcatggaatt ctctctaact gacccgaggc + 6301 tagagccaca caaatgggaa aagtactgtg ttcttgagat aggagacatg ctcctacgga + 6361 ctgcaatagg ccaagtattg aggcccatgt tcctgtatgt gagaaccaat ggaacttcca + 6421 agatcaaaat gaaatggggc atggagatga ggcgatgcct tcttcagtcc cttcaacaaa + 6481 ttgagagcat gattgaggcc gaatcttctg tcaaagagaa ggacatgtcc aaggaattct + 6541 ttgaaaacaa atcagaaaca tggccaattg gagaatcacc caaaggggtg gaggaaggct + 6601 ctattgggaa agtatgcaga acattgctag caaagtctgt gttcaacagc ctatatgcat + 6661 ctcctcaact cgaggggttt tcagctgaat caagaaaatt gcttctcatt gttcaggcac + 6721 ttagggacaa cctggaacct ggaaccttcg atcttggggg gctatatgaa gcaattgagg + 6781 agtgcctgat taacgatccc tgggttttgc ttaatgcatc ttggttcaac tccttcctca + 6841 cacatgcact gaaatagttg tggcaatgct actatttgct atccatactg tccaaaaaag + 6901 taccttgttt ctactgcagg ggttcactct gtcaaaatgg agaacatagt actacttctt + 6961 gcaatagtta gccttgttaa aagtgatcag atttgcattg gttaccatgc aaacaattcg + 7021 acagagcaag ttgacacgat aatggaaaag aacgtcactg ttacacatgc ccaagacata + 7081 ctggaaaaaa cacacaacgg gaagctatgc gacctaaatg gggtgaagcc actgatttta + 7141 aaggactgca gtgtagctgg atggctcctc ggaaacccaa tgtgcgacga attcatcaga + 7201 gtgccggaat ggtcttacat agtggagcgg gctaacccag ctaatgacct ctgttaccca + 7261 gggagcctca atgactatga agaactgaaa cacatgttga gcagaataaa tcattttgag + 7321 aagattcaga tcattcccaa gagttcctgg ccaaatcatg aaacatcact aggggtgagc + 7381 gcagcttgtc cataccaggg agcaccctcc tttttcagaa atgtggtgtg gcttatcaaa + 7441 aagaacgatg catacccaac aataaagata agctacaata atactaatcg ggaagatctc + 7501 ttgatactgt gggggattca tcattccaac aatgcagaag agcagacaaa tctctacaaa + 7561 aacccaatca cctacatttc agttggaaca tcaactttaa accagaggtt ggcaccaaaa + 7621 atagctacta gatcccaagt aaacgggcaa cgtggaagaa tggacttctt ctggacaatc + 7681 ttaaaaccag atgatgcaat ccatttcgag agtaacggaa atttcattgc tccagaatat + 7741 gcatacaaaa ttgttaagaa aggggactcg acaattatga aaagtggagt ggaatatggc + 7801 cactgcaaca ccaaatgtca aaccccagta ggtgcgataa attctagtat gccatttcac + 7861 aacatacatc ctctcaccat tggggaatgc cccaaatacg tgaaatcaaa caagttggtc + 7921 cttgcgactg ggctcagaaa tagtcctcta agagaaaaga gaagaaaann nagaggtctg + 7981 tttggggcga tagcagggtt tatagaggga ggatggcagg gaatggttga tggttggtat + 8041 gggtaccatc atagcaatga gcaggggagt gggtacgctg cggacaaaga atccacccaa + 8101 aaggcaatag atggagttac caataaggtc aactcaatca ttgacaaaat gaacactcaa + 8161 tttgaggcag ttggaaggga gtttaataac ttagaaagga ggatagagaa tttgaacaag + 8221 aaaatggaag acggattcct agatgtctgg acctataatg ctgaacttct agttctcatg + 8281 gaaaacgaga ggactctaga tttccatgat tcaaatgtca agaaccttta cgacaaagtc + 8341 agattacagc ttagggataa tgcaaaggag ctgggtaacg gctgtttcga attctatcac + 8401 aaatgtgata atgaatgtat ggaaagtgtg agaaatggga cgtatgacta ccctcagtat + 8461 tcagaagaag caagattaaa aagagaagaa ataagcggag tgaaattaga atcagtagga + 8521 acttaccaga tactgtcaat ttattcaaca gcggcaagtt ccctagcact ggcaatcatg + 8581 atggctggtc tatctttatg gatgtgctcc aatgggtcgt tacaatgcag aatttgcatt + 8641 tagatttatg agctcagatt gtagttaaaa acaccagcaa aagcagggta gataatcact + 8701 cactgagtga catccacatc atggcgtctc aaggcaccaa acgatcctat gaacaaatgg + 8761 aaactggtgg ggaacgccag aatgccactg aaatcagagc atctgttgga agaatggttg + 8821 gcggaatcgg gagattctac atacagatgt gcactgagct caaactcagt gatcacgaag + 8881 ggaggctgat ccaaaacagc ataaccatag aaaggatggt tctctcggca tttgatgaaa + 8941 ggaggaacaa gtatctggag gaacatccca gtgctggaaa ggatcccaag aagactggag + 9001 gtccaatcta caggaggaga gatggcaaat ggatgagaga gttgatcctc tacgacaaag + 9061 aagagatcag aagaatttgg cgtcaagcta ataatggaga ggatgcaact gctggtctca + 9121 ctcatttgat gatttggcat tccaatctga atgatgccac ataccagaga acaagggcac + 9181 ttgtgcgtac tggaatggat cctaggatgt gctcactgat gcaaggctca accctcccta + 9241 ggagatccgg ggctgctgga gcggcagtga aaggagttgg aacaatggtg atggaattga + 9301 ttcgaatgat caaacgagga atcaatgatc ggaatttctg gagaggtgaa aacggacgga + 9361 gaaccaggat tgcctacgag agaatgtgca acatcctcaa gggaaagttc caaacagcag + 9421 cacaacgagc aatgatggac caagtgaggg aaagccggaa tcctgggaat gctgaaattg + 9481 aagatctcat ctttctcgca cgatctgctc tcatcctgag gggatcagtg gctcataagt + 9541 cctgtctgcc tgcttgcgtg tatggacttg ctgtagccag tggatatgac tttgaaagag + 9601 agggatactc tctagtcgga attgatcctt tccgtctgct ccagaacagt caagttttca + 9661 gtctcatcag accgaatgaa aatccagctc acaaaagtca gctggtatgg atggcatgcc + 9721 actctgcggc atttgaggat ctgagagtgt caagcttcat cagaggaaca agagtagtcc + 9781 caagaggaca actgtccacc agaggagttc agattgcttc aaatgaaaac atggagacaa + 9841 tggattccag tactcttgaa ctgaggagca gatactgggc tataagaaca agaagtggag + 9901 gaaacaccaa ccaacagaga gcatctgcag gacaaatcag cgtacagccc acattctctg + 9961 tgcagagaaa cctcccattc gagagagcaa ccatcatggc agcatttacg ggaaacactg + 10021 aaggcagaac ttcagacatg agaactgaga tcataaggat gatggaaaat gccagacctg + 10081 aagatgtgtc tttccagggg cggggagtct tcgagctctc ggacgaaaag gcaacgaacc + 10141 cgatcgtgcc ttcctttgac atgaacaatg aaggatctta tttcttcgga gacaatgcag + 10201 aggagtatga caattaaaga aaaataccct tgtttctact agcaaaagca ggagttcaaa + 10261 atgaatccaa atcaaaagat aacaaccatt ggatcaatct gtatggtaat tgggatagtc + 10321 agcttgatgc tgcaaattgg gaacataatc tcaatatggg ttagccattc aatccaaaca + 10381 gggaatcaat accagcctga accatgcaat caaagcatca ttacctatga gaacaacacc + 10441 tgggtaaatc agacgtatat caacatcagc aataccaatt ttcttgctga gcaggctgtt + 10501 acttcggtaa cattagcggg caattcatct ctttgcccta ttagtgggtg ggcaatatac + 10561 agtaaggaca acggtataag aattgggtct aagggggatg tgtttgttat aagagaacca + 10621 ttcatctcat gctcccactt ggaatgcaga acctttttcc tgacccaggg agctctgctg + 10681 aatgacaaac attctaatgg gacagttaag gatagaagcc cttatagaac tttgatgagt + 10741 tgtcccgtgg gtgaggctcc ttccccgtac aattcaagat ttgagtctgt tgcttggtcg + 10801 gcaagtgctt gtcatgatgg catcagttgg ttgacaatcg gtatttctgg tccagacaat + 10861 ggagctgtgg ctgtattgaa gtacaatggc ataataacgg atactatcaa gagttggaga + 10921 aacaacattt tgagaactca agaatctgaa tgtgcttgcg taaatggctc ctgcttcacc + 10981 gtaatgactg atggaccaag caatgggcag gcctcatata aaatcttcaa gatagagaaa + 11041 gggaaagttg tcaaatcagt tgaaatgaat gcccctaatt accactacga ggaatgctcc + 11101 tgttatcctg atgcgggtga tattatgtgt gtgtgcaggg acaattggca tggctcgaac + 11161 cggccgtggg tatcttttaa tcaaaatctg gagtatcaaa taggatatat atgcagtggg + 11221 attttcgggg acaatccccg ccccaatgat ggaacaggca gttgcagtcc aatgccctct + 11281 aatggggcat atggggtaaa agggttttca tttaagtacg gtaatggggt ttggatcgga + 11341 agaacaaaaa gcactagttc cagaagcggc tttgagatga tttgggatcc gaatgggtgg + 11401 actgagacgg acagtagttt ctcagtgaag caagacattg tagaaataac tgactggtca + 11461 ggatatagtg ggagttttgt ccagcatcca gaactgacag gattagattg catgaggcct + 11521 tgtttctggg ttgagctaat tagagggagg cccaaagaga atacaatttg gactagcggg + 11581 agcagcatat ccttttgtgg tgtaaatagt gacactgtgg gttggtcttg gccagacggt + 11641 gctgagttgc cattcaccat tgacaagtag tttgttcaaa aaactccttg tttctactag + 11701 caaaagcagg tagatattga aagatgagtc ttctaaccga ggtcgaaacg tacgttctct + 11761 ctatcgtccc gtcgggcccc ctcaaagccg agatcgcgca gagacttgaa gatgtctttg + 11821 cagggaagaa caccgatctt gaggctctca tggaatggct aaagacaaga ccaatcctgt + 11881 cacctctgac taaggggatt ttgggatttg tgttcacgct caccgtgccc agtgagcgag + 11941 gactgcagcg tagacgcttt gtccaaagtg ccctaagtgg aactggagac ccaaacaaca + 12001 tggacagagc agtcaagttg tacaggaaac tgaagagaga gataacattc catggggcta + 12061 aagaagttgc actcagttac tcaaccggtg cacttgccag ttgtatgggt ctcatataca + 12121 acaggatggg gacggtgacc gcagaagtgg cattgggcct agtgtgtgcc acctgtgaac + 12181 agattgctga ttcacagcat cggtctcaca gacagatagc taccaccacc aacccactga + 12241 tcagacatga aaacagaatg gttttggcca gtactacagc taaggctatg gagcagatgg + 12301 ctggatcgag tgagcaagca gtggaagcca tggaggttgc tagtcaggct aggcagatgg + 12361 tgcaggcgat gaggaccatt ggaactcatc ctagctccag taccggtctg agagatgatc + 12421 tccttgaaaa tttgcaggcc taccaaaaac ggatgggagt gcaactgcag cgattcaagt + 12481 gatcctctcg ttattgccgc aagtatcatt gggatcttgc acttgatatt gtggattctt + 12541 gatcgccttt tcttcaaatg cgtttatcgt cgccttaaat acggtttgaa aggagggcct + 12601 tctacggaag gagtacctga gtccatgagg gaagagtacc ggcaggaaca gcagagtgct + 12661 gtggatgttg acaatggtca ttttgtcaac atagagctgg agtaaaaaac taccttgttt + 12721 ctactgtgac aaaaacataa tggattccaa cactgtgtta agctttcagg tagactgctt + 12781 tctttggcat gtccgcaaac gatttgcaga ccaagaactg ggtgatgccc cattccttga + 12841 ccggctccgc cgagaccaga agtctctaag aggaagaggc agcactcttg gtctggacat + 12901 cgagacggcc actcgtgctg ggaagcagat agtggagagg attctggagg aagaatccga + 12961 cgaggcactc aaaatgacta ttgcctctgt gcctgctcca cgctacctaa ctgacatgac + 13021 tcttgaagag atgtcaagag actggttcat gctcatgccc aagcaaaaag tggcaggctc + 13081 cctcagtatc agaatggacc aggcgattat ggataagaac atcatactga aggcaaactt + 13141 cagtgtgatc ttcaatcggc tggagacact aatactactc agagctttca ctgaagaggg + 13201 agcaattgtc ggcgaaattt caccattgcc ttctcttcca ggacatactg atgaggatgt + 13261 caaaaatgca attggggtcc tcatcggagg acttgaatgg aatgataaca cagttcgagt + 13321 ctctgaaact ttacagagat tcgcttggag aagcagtaat gaggatggga gacctccact + 13381 ccctccaaag cagaaacgga aaatggagag gacaattgag tcagaagttt gaagaaataa + 13441 ggtggctgat tgaagaagtg cgacacagac taaagatcac agaaaatagt tttgaacaaa + 13501 taacatttat gcaagcctta caactactgc ttgaagtgga gcaagagata agaactttct + 13561 cgtttcagct tatttaataa taaaaaacac +//