淀粉样变性病实用课件

NCBI FieldGuideGenome ResourcesLocusLinkLocusLinkLocusLinkLocusLinkGene databaseGene databaseGene databaseGene databaseUniGeneUniGeneUniGeneUniGeneTrace ArchiveTrace ArchiveTrace ArchiveTrace ArchiveMap ViewerMap ViewerMap ViewerMap ViewerHomologeneHomologeneHomologeneHomologeneGenome ResourcesLocusLinkGene 1NCBI FieldGuideGenomic BiologyGenomic Biology2NCBI FieldGuide淀粉样变性病实用课件3NCBI FieldGuide淀粉样变性病实用课件4NCBI FieldGuide淀粉样变性病实用课件5NCBI FieldGuideGenome Projects:microbGenome Projects:microb6NCBI FieldGuide淀粉样变性病实用课件7NCBI FieldGuide=scale for scoring systemNegative for less likely substitutionsOther purine nucleotide metabolizing enzymes not found by ordinary BLASTMus musculusOrganismG 0-2 0-1-3-2-2 6nucleotide,protein and translations(blastn,blastp,blastx,tblastn,tblastx)environmental samplefilter,e.GTACTGGACATSbjct 287 LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEI 333catalytic loopG 0-2 0-1-3-2-2 6Word size=3(default)refseq_genomic=NC_*,NG_*Entrez GeneEntrez linksDrosophila melanogaster CG18582-PA(mbt)mRNA,(3244 bp)Why Do We Need Sequence Similarity Searching?UniGene Cluster Hs.Service Addresses95351:seqs=scale for scoring system8NCBI FieldGuideGenome ResourcesLocusLinkLocusLinkLocusLinkLocusLinkGene databaseGene databaseGene databaseGene databaseUniGeneUniGeneUniGeneUniGeneTrace ArchiveTrace ArchiveTrace ArchiveTrace ArchiveMap ViewerMap ViewerMap ViewerMap ViewerHomologeneHomologeneHomologeneHomologeneGenome ResourcesLocusLinkGene 9NCBI FieldGuide451 N -4-3 8-1-5-2-2-3-1-6-6-2-4-5-4-1-2-6-4-5Nucleotide only:-e 10000-v 2000ATCACCATGAAGTGGCTGAAGGATAAGCAGCCAATGGATGCCAAGGAGTTCGAACCTAAAGACGTATTGCCCAATGGGGATGGGACBasic Local Alignment Search Toolbiomol mrnaPropertiesSbjct 287 LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEI 333RESULTS:Initial BLASTPSpecies Number of genesW=11,t=18,non-coding:111010010110010111Make lookup table of“words”for querySsc_UniGenevs DrosophilaWhitehead-RHGTQITVEDLFYNIATRRKALKNSerine/Threonine protein kinasesGTACTGGACATGGACCCTACAGGAALKSEDDLLNFPSVEHVTSVVLKRMICNALIDRPNTLFVFDDVVQEETIRWAQELRLRCLVTTRDVEIA single query interface to SequencesSequences-RefSeqs-RefSeqs-GenBank-GenBank-Homologene-HomologeneMaps MapViewerMaps MapViewerEntrez linksEntrez linksLocusLinkLocusLink will be replaced by Entrez Gene on MARCH 1,2005.Check Gene FAQ for current information.451 N -4-3 8-1-5-2-2-10NCBI FieldGuideEntrez GeneLocusLinkA single query interface to SequencesSequences -RefSeqs -RefSeqs -GenBank -GenBank -Homologene -HomologeneMaps MapViewerMaps MapViewerEntrez linksEntrez linksEntrez Gene More organisms-all RefSeq genomes Entrez integrationEntrez GeneLocusLinkA single q11NCBI FieldGuideGsnsym淀粉样变性病Gsnsym淀粉样变性病12NCBI FieldGuide淀粉样变性病实用课件13NCBI FieldGuideGlobal Entrez:NADH2nadh247Global Entrez:NADH2nadh24714NCBI FieldGuideEntrez Gene:NADH226 recordsEntrez Gene:NADH226 records15NCBI FieldGuideGene Record for Pongo NADH2Homo sapiensGene Record for Pongo NADH2Hom16NCBI FieldGuideDisplay Exons/Introns:Gene TableDisplay Exons/Introns:Gene T17NCBI FieldGuideGene TableGene Table18NCBI FieldGuideA Record With More Data:Human HFE血色沉着病A Record With More Data:Human19NCBI FieldGuideGene Graphic LinksNM_NM_NP_NP_Gene Graphic LinksNM_NP_20NCBI FieldGuideIntrons/Exons:Gene Tablelinks to sequenceIntrons/Exons:Gene Tablelin21NCBI FieldGuideGga_UniGeneFinds best local alignmentsGTACTGGACAT441 A 3-2 1-2 0-1 0 1-2-2-2 0-1-2 3 1 0-3-3 0H-2 0 1-1-3 0 0-2 8The New HomologeneJust below threshold,anotherSsc_UniGenesp|P27476|NSR1_YEAST NUCLEAR LOCALIZATION SEQUENCE BINDING PROTEIN(P67)Other BLAST AlgorithmsYLS HFLIdentity matrix450 K 0 3 0 1-5 0 0-4-1-4-3 4-3-2 2 1-1-5-4-4Trace ArchiveStrongylocentrotus purpuratus TracesMegaBLAST=“No significant similarity found.Nucleotide only:gray line=same database hitProvides statistical significancecatalytic loopBasic Local Alignment Search ToolA Record With More Data:Human HFEGga_UniGeneA Record With More 22NCBI FieldGuideEntrez SNPhfegene name AND humanorgn 52血色沉着病Entrez SNPhfegene name AND h23NCBI FieldGuideLinking to SNP染色体定位基因定位序列定位Linking to SNP染色体定位基因定位序列定位24NCBI FieldGuideSNP in StructureSNP in Structure25NCBI FieldGuideLink to OMIMLink to OMIM26NCBI FieldGuideVariants in OMIMVariants in OMIM27NCBI FieldGuideGenome ResourcesLocusLinkLocusLinkLocusLinkLocusLinkGene databaseGene databaseGene databaseGene databaseTrace ArchiveTrace ArchiveTrace ArchiveTrace ArchiveMap ViewerMap ViewerMap ViewerMap ViewerHomologeneHomologeneHomologeneHomologeneUniGeneUniGeneGenome ResourcesLocusLinkGene 28NCBI FieldGuideACATGGACCCTGTQITVEDLFYNI=scale for scoring systemgi|113340|sp|P03958|ADA_MOUSE ADENOSINE DEAMINASE(ADENOSINE435 K -1 0 0-1-2 3 0 3 0-2-2 1-1-1-1-1-1-1-1-2The New HomologeneBLOSUM62-default matrix for BLASTTrade-off:sensitivity vs speedSsc_UniGeneRAG1 HomologeneDiscontiguous(Cross-species)MegaBLASTDisplay Exons/Introns:Gene TableMammaliaOrganismbiomol mrnaPropertiesUniGene Cluster Hs.sp|P27476|NSR1_YEAST NUCLEAR LOCALIZATION SEQUENCE BINDING PROTEIN(P67)GTQITVEDLFYNILimit by taxonEntrez Gene:NADH2Mus musculusOrganismGene-oriented clusters of expressed sequences Automatic clustering using MegaBlast Each cluster represents a unique gene Informed by genome hits Information on tissue types and map locations Useful for gene discovery and selection of mapping reagentsUniGeneACATGGACCCTGene-oriented clust29NCBI FieldGuideSeq1:1 W-HEREISWALTERNOW 16No longer UniGene basedoptimized for large batch searchesL-1-2-3-4-1-2-3-4-3 2 4GAxxxxGKSTnucleotide metabolism proteinsCheck to add to PSSMcatalytic loopcheck primer specificity in silicoIdentity matrixWord size can only be 2 or 3-f 11=blastp default W=12,t=21,coding:100101101101100101101BLOSUM62-default matrix for BLASTGlobal vs Local Alignment452 I -3-5-5-6 0-5-5-6-5 6 2-5 2-2-5-4-3-5-3 3GTQITVEDLFYNIDerived from observation;small dataset of alignmentsStandard BLASTG 0-2 0-1-3-2-2 6A Cluster of ESTsquery5 EST hits3 EST hitsSeq1:1 W-HERE30NCBI FieldGuideUnigeneUnigene31NCBI FieldGuideUniGene CollectionsUniGene Collections32NCBI FieldGuideExample UniGene ClusterExample UniGene Cluster33NCBI FieldGuideHistogram of cluster sizes for UniGene Hs build 177Histogram of cluster sizes for34NCBI FieldGuideX 0-1-1-1-2-1-1-1-1-1-1-1-1-1-2 0 0-2-1-1-1458 L -3-1 0-3 0-3-2 3-4-2 3 0 1 1-2-2-3 5-1-3Discontiguous MegablastLTV,MTV,ISV,LSV,etc.A R N D C Q E G H I L K M F P S T W Y VJust below threshold,anotherHuman Genome BLAST:Resultsvery sensitive protein searchDiscontiguous megaBLAST=Gene databaseGeneral HelpProtein vs DNA translation tblastn446 D -4-4-1 8-6-2 0-3-3-5-6-3-5-6-4-2-3-7-5-5H-2 0 1-1-3 0 0-2 8441 A 3-2 1-2 0-1 0 1-2-2-2 0-1-2 3 1 0-3-3 0=scale for scoring systemHomolgene:RAG1W=word size;#matches in templateGTQITVEDLFYNISequence View (sv)UniGene Cluster Hs.95351X 0-1-1-1-2-1-1-1-1-35NCBI FieldGuideUniGene Cluster Hs.95351UniGene Cluster Hs.9535136NCBI FieldGuideUniGene Cluster Hs.95351:expressionUniGene Cluster Hs.95351:expr37NCBI FieldGuideUniGene Cluster Hs.95351:seqsUniGene Cluster Hs.95351:seqs38NCBI FieldGuideDownload sequencesweb pageftp siteDownload sequencesweb pageftp 39NCBI FieldGuideGenome ResourcesLocusLinkLocusLinkLocusLinkLocusLinkGene databaseGene databaseGene databaseGene databaseUniGeneUniGeneUniGeneUniGeneTrace ArchiveTrace ArchiveTrace ArchiveTrace ArchiveMap ViewerMap ViewerMap ViewerMap ViewerHomologeneHomologeneHomologeneHomologeneGenome ResourcesLocusLinkGene 40NCBI FieldGuideThe New HomologeneAutomated detection of homologs among the annotated genes of completely sequenced eukaryotic genomes.No longer UniGene basedProtein similarities firstGuided by taxonomic treeIncludes orthologs and paralogsThe New HomologeneAutomated de41NCBI FieldGuide Orthologs 和 Paralogs 是同源序列的两种类型。Orthologs（垂直同源基因）是指来自于不同物种的由垂直家系（物种形成）进化而来的蛋白，并且典型的保留与原始蛋白有相同的功能。Paralogs（平行同源基因）是那些在一定物种中的来源于基因复制的蛋白，可能会进化出新的与原来有关的功能。请参考文献获得更多的信息。Orthologs 和 Paralogs42NCBI FieldGuidegene duplicationParalogs vs Orthologsearly globin geneA-chain gene B-chain genefrog A chick A mouse Amouse B chick B frog Bparalogsorthologs orthologsgene duplicationParalogs vs Or43NCBI FieldGuideThe New Homologene Homologene Build 37.2Species Number of genes input grouped groupsThe New Homologene 44NCBI FieldGuideGlobal vs Local AlignmentHuman Genome BLAST:Resultsreverse primerBasic Local Alignment Search ToolSeq1:1 W-HEREISWALTERNOW 16GTQITVEDLFYNIreverse primerAll combinations(DNA/Protein)query and database.“hemochromatosis”Sbjct 287 LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEI 333-Genetic Maps-W=word size;#matches in templateStandard BLASTMegaBLAST vs Discontiguous MegaBLASTProtein BLAST requires two neighboring matches within 40 aaX 0-1-1-1-2-1-1-1-1-1-1-1-1-1-2 0 0-2-1-1-1-b 2000alignmentsA Record With More Data:Human HFEWayne Matten456 N 1 1 3 0-4-1 1 0-3-4-4 3-2-5-2 2-2-5-4-4RAG1 Homologenerag112recombination activating gene Global vs Local AlignmentRAG1 45NCBI FieldGuideRAG1 HomolgeneRAG1Amniota RAG1 HomolgeneRAG1Amniota 46NCBI FieldGuideHomolgene:RAG1Homolgene:RAG147NCBI FieldGuide淀粉样变性病实用课件48NCBI FieldGuideHomolgene:RAG1Homolgene:RAG149NCBI FieldGuideGenome ResourcesLocusLinkLocusLinkLocusLinkLocusLinkGene databaseGene databaseGene databaseGene databaseUniGeneUniGeneUniGeneUniGeneTrace ArchiveTrace ArchiveTrace ArchiveTrace ArchiveMap ViewerMap ViewerMap ViewerMap ViewerHomologeneHomologeneHomologeneHomologeneGenome ResourcesLocusLinkGene 50NCBI FieldGuide淀粉样变性病实用课件51NCBI FieldGuide淀粉样变性病实用课件52NCBI FieldGuide淀粉样变性病实用课件53NCBI FieldGuideMapViewerMapViewer54NCBI FieldGuideList ViewList View55NCBI FieldGuideMore info:Ex:BLASTNMammaliaOrganismGenome ResourcesMegaBLAST=“No significant similarity found.Genome ResourcesStandford-G3441 A 3-2 1-2 0-1 0 1-2-2-2 0-1-2 3 1 0-3-3 0|raw score=19-9=10Genome ResourcesProvides statistical significanceDisplay Exons/Introns:Gene TableMus musculusOrganismK=scale for search spacegov/gene/DATA/gene_info.Species Number of genesUniGene Cluster Hs.A+1 3 3-3Human MapVieweradar腺甙脱氨酶More info:Human MapVieweradar腺56NCBI FieldGuideMapViewer:Human ADAR4MapViewer:Human ADAR457NCBI FieldGuideMV Hs ADAR3 UTR5 UTRMV Hs ADAR3 UTR5 UTR58NCBI FieldGuideMaps&Options-Sequence mapsSequence maps-Ab initioAssemblyRepeatsBES_CloneCloneNCI_CloneContigComponentCpG islanddbSNP haplotypeFosmidGenBank_DNAGenePhenotypeSAGE_TagSTSTCAG_RNATranscript(RNA)Hs_UniGeneHs_EST-Cytogenetic mapsCytogenetic maps-IdeogramFISH CloneGene_CytogeneticMitelman BreakpointMorbid/Disease-Genetic Maps-deCODEGenethonMarshfield-RH maps-GeneMap99-G3GeneMap99-GB4NCBI RHStandford-G3TNGWhitehead-RHWhitehead-YACMm_UniGeneMm_ESTRn_UniGeneRn_ESTSsc_UniGeneSsc_ESTBt_UniGeneBt_ESTGga_UniGeneGga_ESTVariationMaps&Options=SNPMaps&Options-Sequence maps-59NCBI FieldGuideMapViewerUniGeneComponentRepeatsGeneMapViewerUniGeneComponentRepea60NCBI FieldGuideMaster map:repeatsMaster map:repeats61NCBI FieldGuideGenePhenotypeVariationGenePhenotypeVariation62NCBI FieldGuideW=12,t=18,coding:101101101101101101GAxxxxGKSTCAGGTAGCAAGCTTGCATGTCATrace Archiverecombination activating geneUniGene Cluster Hs.446 D -4-4-1 8-6-2 0-3-3-5-6-3-5-6-4-2-3-7-5-5DNA translation vs DNA translation tblastxK=scale for search spaceGenomic Biology2 bits(92),Expect=0.Sbjct 287 LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEI 333refseq_genomic=NC_*,NG_*441 A 3-2 1-2 0-1 0 1-2-2-2 0-1-2 3 1 0-3-3 0BLAST statisticsray finned fishesOrganismParticularly useful for nucleotide sequences without440 P -2-2-2-2-3-2-2-2-2-1-2-1 0-3 7-1-2-3-1-1CATGCTTAATTAll combinations(DNA/Protein)query and database.PAM250 widely usedMaps&OptionsMaps&OptionsW=12,t=18,coding:1011063NCBI FieldGuideGenome ResourcesLocusLinkLocusLinkLocusLinkLocusLinkGene databaseGene databaseGene databaseGene databaseUniGeneUniGeneUniGeneUniGeneTrace ArchiveTrace ArchiveTrace ArchiveTrace ArchiveMap ViewerMap ViewerMap ViewerMap ViewerHomologeneHomologeneHomologeneHomologeneGenome ResourcesLocusLinkGene 64NCBI FieldGuide淀粉样变性病实用课件65NCBI FieldGuide淀粉样变性病实用课件66NCBI FieldGuideStrongylocentrotus purpuratus TracesStrongylocentrotus purpuratus 67NCBI FieldGuideBLASTBasic Local Alignment Search ToolBLASTBasic Local Alignment Sea68NCBI FieldGuideWeb AccessBLASTVASTEntrezTextSequenceStructureWeb AccessBLASTVASTEntrezTextS69NCBI FieldGuide淀粉样变性病实用课件70NCBI FieldGuideBasic Local Alignment Search Tool Why use sequence similarity?BLAST algorithm BLAST statistics BLAST output ExamplesBasic Local Alignment Search T71NCBI FieldGuideWhy Do We Need Sequence Similarity Searching?To identify and annotate sequencesTo evaluate evolutionary relationshipsOther:model genomic structure(e.g.,Spidey)check primer specificity in silico:NCBIs toolWhy Do We Need Sequence Simil72NCBI FieldGuideBLAST Website StatsBLAST Website Stats73NCBI FieldGuideGlobal vs Local AlignmentSeq 1Seq 2Seq 1Seq 2Global alignmentLocal alignmentGlobal vs Local AlignmentSeq 174NCBI FieldGuideGlobal vs Local AlignmentSeq1:WHEREISWALTERNOW (16aa)Seq2:HEWASHEREBUTNOWISHERE(21aa)GlobalSeq1:1 W-HEREISWALTERNOW 16 W HERE Seq2:1 HEWASHEREBUTNOWISHERE 21LocalSeq1:1 W-HERE 5 Seq1:1 W-HERE 5 W HERE W HERESeq2:3 WASHERE 9 Seq2:15 WISHERE 21Global vs Local AlignmentSeq1:75NCBI FieldGuideThe Flavors of BLASTStandard BLASTtraditional“contiguous”word hitposition independent scoring nucleotide,protein and translations(blastn,blastp,blastx,tblastn,tblastx)Megablastoptimized for large batch searchescan use discontiguous wordsPSI-BLASTconstructs PSSMs automatically;uses as queryvery sensitive protein searchRPS BLASTsearches a database of PSSMstool for conserved domain searchesThe Flavors of BLASTStandard B76NCBI FieldGuideWidely used similarity search toolHeuristic approach based on Smith Waterman algorithmFinds best local alignmentsProvides statistical significanceAll combinations(DNA/Protein)query and database.DNA vs DNA blastnDNA translation vs Protein blastxProtein vs Protein blastpProtein vs DNA translation tblastnDNA translation vs DNA translation tblastx www,standalone,and network clientsBasic Local Alignment Search ToolWidely used similarity search 77NCBI FieldGuideTranslated BLASTQueryQueryDatabaseDatabaseProgramProgramNPucleotideroteinNNNNPPblastxtblastntblastxPPPPPPPPPPPPPPPPPPPPPPPPParticularly useful for nucleotide sequences withoutprotein annotations,such as ESTs or genomic DNATranslated BLASTQueryDatabaseP78NCBI FieldGuideHow BLAST WorksMake lookup table of“words”for queryScan database for hitsUngapped extensions of hits(initial HSPs)Gapped extensions(no traceback)Gapped extensions(traceback;alignment details)How BLAST WorksMake lookup tab79NCBI FieldGuideNucleotide WordsGTACTGGACATGGACCCTACAGGAAQuery:GTACTGGACAT TACTGGACATG ACTGGACATGG CTGGACATGGA TGGACATGGAC GGACATGGACC GACATGGACCC ACATGGACCCTMake a lookuptable of words11-mer.828megablast711blastnminimumdefaultWORD SIZENucleotide WordsGTACTGGACATGGA80NCBI FieldGuideProtein WordsGTQITVEDLFYNIATRRKALKNQuery:Neighborhood WordsLTV,MTV,ISV,LSV,etc.GTQ TQI QIT ITV TVE VED EDL DLF .Make a lookuptable of wordsWord size=3(default)Word size can only be 2 or 3-f 11=blastp default Protein WordsGTQITVEDLFYNIATRR81NCBI FieldGuideMinimum Requirements for a Hit Nucleotide BLAST requires one exact match Protein BLAST requires two neighboring matches within 40 aaGTQITVEDLFYNI SEI YYNATCGCCATGCTTAATTGGGCTT CATGCTTAATT neighborhood wordsone exact matchtwo matches-A 40=blastp default Minimum Requirements for a Hit82NCBI FieldGuideBLASTP Summary YLS HFLSbjct 287 LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEI 333 Query 1 IETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESI 47Gapped extension with trace backGapped extension with trace backQuery 1 IETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESI-LEV 50 +E YA YL K F+YLSL+SP+DVNVHP+K VHFL+I+Sbjct 287 LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEIATSI 337 Final HSPFinal HSP +E YA YL K F+L+SP+DVNVHP+K V +I High-scoring pair(HSP)High-scoring pair(HSP)HFL 18HFV 15 HFS 14HWL 13NFL 13DFL 12HWV 10etc YLS 15YLT 12 YVS 12YIT 10etc Neighborhood Neighborhood wordswordsNeighborhood Neighborhood score thresholdscore thresholdT(-f)=11T(-f)=11Query:IETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESILEVexample query wordsexample query wordsBLASTP Summary 83NCBI FieldGuideScoring Systems-Nucleotides A G C TA+1 3 3-3G 3+1 3-3C 3 3+1-3T 3 3 3+1Identity matrixCAGGTAGCAAGCTTGCATGTCA|raw score=19-9=10CACGTAGCAAGCTTG-GTGTCA-r 1-q-3 Scoring Systems-Nucleotides 84NCBI FieldGuideScoring Systems-ProteinsPosition Independent MatricesPAM Matrices(Percent Accepted Mutation)Derived from observation;small dataset of alignments Implicit model of evolution All calculated from PAM1 PAM250 widely usedBLOSUM Matrices(BLOck SUbstitution Matrices)Derived from observation;large dataset of highly conserved blocks Each matrix derived separately from blocks with a defined percent identity cutoff BLOSUM62-default matrix for BLASTPosition Specific Score Matrices(PSSMs)PSI-and RPS-BLASTScoring Systems-ProteinsPosi85NCBI FieldGuideA 4R-1 5 N-2 0 6D-2-2 1 6C 0-3-3-3 9Q-1 1 0 0-3 5E-1 0 0 2-4 2 5G 0-2 0-1-3-2-2 6H-2 0 1-1-3 0 0-2 8I-1-3-3-3-1-3-3-4-3 4 L-1-2-3-4-1-2-3-4-3 2 4K-1 2 0-1-3 1 1-2-1-3-2 5M-1-1-2-3-1 0-2-3-2 1 2-1 5F-2-3-3-3-2-3-3-3-1 0 0-3 0 6P-1-2-2-1-3-1-1-2-2-3-3-1-2-4 7S 1-1 1 0-1 0 0 0-1-2-2 0-1-2-1 4T 0-1 0-1-1-1-1-2-2-1-1-1-1-2-1 1 5W-3-3-4-4-2-2-3-2-2-3-2-3-1 1-4-3-2 11Y-2-2-2-3-2-1-2-3 2-1-1-2-1 3-3-2-2 2 7V 0-3-3-3-1-2-2-3-3 3 1-2 1-1-2-2 0-3-1 4X 0-1-1-1-2-1-1-1-1-1-1-1-1-1-2 0 0-2-1-1-1 A R N D C Q E G H I L K M F P S T W Y V XBLOSUM62DFNegative for less likely substitutionsDYFPositive for more likely substitutionsA 4BLOSUM62DFNegative for les86NCBI FieldGuidePosition-Specific Score MatrixDAF-1Serine/Threonine protein kinases catalytic loop174PSSM scores54Position-Specific Score Matrix87NCBI FieldGuide A R N D C Q E G H I L K M F P S T W Y V 435 K -1 0 0-1-2 3 0 3 0-2-2 1-1-1-1-1-1-1-1-2 436 E 0 1 0 2-1 0 2-1 0-1-1 0 0 0-1 0 0-1-1-1 437 S 0 0-1 0 1 1 0 1 1 0-1 0 0 0 2 0-1-1 0-1 438 N -1 0-1-1 1 0-1 3 3-1-1 1-1 0 0-1-1 1 1-1 439 K -2 1 1-1-2 0-1-2-2-1-2 5 1-2-2-1-1-2-2-1 440 P -2-2-2-2-3-2-2-2-2-1-2-1 0-3 7-1-2-3-1-1 441 A 3-2 1-2 0-1 0 1-2-2-2 0-1-2 3 1 0-3-3 0 442 M -3-4-4-4-3-4-4-5-4 7 0-4 1 0-4-4-2-4-1 2 443 A 4-4-4-4 0-4-4-3-4 4-1-4-2-3-4-1-2-4-3 4 444 H -4-2-1-3-5-2-2-4 10-6-5-3-4-3-2-3-4-5 0-5 445 R -4 8-3-4 0-1-2-3-2-5-4 0-3-2-4-3-3 0-4-5 446 D -4-4-1 8-6-2 0-3-3-5-6-3-5-6-4-2-3-7-5-5 447 I -4-5-6-6-3-4-5-6-5 3 5-5 1 1-5-5-3-4-3 1 448 K 0 0 1-3-5-1-1-3-3-5-5 7-4-5-3-1-2-5-4-4 449 S 0-3-2-3 0-2-2-3-3-4-4-2-4-5 2 6 2-5-4-4 450 K 0 3 0 1-5 0 0-4-1-4-3 4-3-2 2 1-1-5-4-4 451 N -4-3 8-1-5-2-2-3-1-6-6-2-4-5-4-1-2-6-4-5 452 I -3-5-5-6 0-5-5-6-5 6 2-5 2-2-5-4-3-5-3 3 453 M -4-4-6-6-3-4-5-6-5 0 6-5 1 0-5-4-3-4-3 0 454 V -3-3-5-6-3-4-5-6-5 3 3-4 2-2-5-4-3-5-3 5 455 K -2 1 1 4-5 0-1-2 1-4-2 4-3-2-3 0-1-5-2-3 456 N 1 1 3 0-4-1 1 0-3-4-4 3-2-5-2 2-2-5-4-4 457 D -3-2 5 5-1-1 1-1 0-5-4 0-2-5-1 0-2-6-4-5 458 L -3-1 0-3 0-3-2 3-4-2 3 0 1 1-2-2-3 5-1-3Position-Specific Score Matrixcatalytic loop./blastpgp-i NP_499868.2-d nr-j 3-Q NP_499868.pssm A R N D C Q E88NCBI FieldGuideLocal Alignment StatisticsHigh scores of local alignments between two random sequencesfollow the Extreme Value DistributionScore(S)Alignments(applies to ungapped alignments)E=Kmne-S or E=mn2-SK=scale for search space =scale for scoring system S=bitscore=(S-lnK)/ln2Expect ValueExpect ValueE=number of database hits you expect to find by chance,Syour scoreexpected number of random hitsMore info:Local Alignment StatisticsHigh89NCBI FieldGuideAdvanced BLAST Options:NucleotideExample Entrez Queriesnucleotide allFilter NOT mammaliaOrganismgreen plantsOrganismbiomol mrnaPropertiesgbdiv estProperties AND ratorganismOther Advancede 10000 expect value-v 2000 descriptions-b 2000 alignmentsAdvanced BLAST Options:Nucleo90NCBI FieldGuideAdvanced BLAST Options:ProteinMatrix SelectionPAM30-most stringentBLOSUM45-least stringentExample Entrez Queriesproteins allFilter NOT mammaliaOrganismgreen plantsOrganismsrcdb refseqPropertiesOther Advancede 10000 expect value-v 2000 descriptions-b 2000 alignmentsLimit by taxonMus musculusOrganismMammaliaOrganismViridiplantaeOrganismAdvanced BLAST