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AFieldGuide
part2
GenomeresourcesSequencesimilarityApr,2007ShandongUniversityGenomeResourcesLocusLinkGenedatabaseUniGeneTraceArchiveMapViewerHomologeneGenomicBiologyGenomeProjects:microbGenomeResourcesLocusLinkGenedatabaseUniGeneTraceArchiveMapViewerHomologeneAsinglequeryinterfaceto…
Sequences -RefSeqs -GenBank -HomologeneMaps–MapViewerEntrezlinksLocusLinkLocusLinkwillbereplacedbyEntrezGeneonMARCH1,2005.CheckGeneFAQforcurrentinformation.EntrezGeneLocusLinkAsinglequeryinterfaceto…
Sequences-RefSeqs-GenBank-HomologeneMaps–MapViewer
EntrezlinksEntrezGene
Moreorganisms-allRefSeqgenomesEntrezintegrationGsn[sym]淀粉樣變性病GlobalEntrez:NADH2nadh247EntrezGene:NADH226recordsGeneRecordforPongoNADH2HomosapiensDisplayExons/Introns:GeneTableGeneTableARecordWithMoreData:HumanHFE血色沉著病GeneGraphicLinksNM_NP_Introns/Exons:GeneTablelinkstosequenceARecordWithMoreData:HumanHFEEntrezSNPhfe[genename]ANDhuman[orgn]52血色沉著病LinkingtoSNP染色體定位基因定位序列定位SNPinStructureLinktoOMIMVariantsinOMIMGenomeResourcesLocusLinkGenedatabaseTraceArchiveMapViewerHomologeneUniGeneGene-orientedclustersofexpressedsequences
AutomaticclusteringusingMegaBlastEachclusterrepresentsauniquegeneInformedbygenomehitsInformationontissuetypesandmaplocationsUsefulforgenediscoveryandselectionofmapping reagentsUniGeneAClusterofESTsquery5’ESThits3’ESThitsUnigeneUniGeneCollectionsExampleUniGeneClusterHistogramofclustersizesforUniGeneHsbuild177UniGeneClusterHs.95351UniGeneClusterHs.95351UniGeneClusterHs.95351:expressionUniGeneClusterHs.95351:seqsDownloadsequenceswebpageftpsiteGenomeResourcesLocusLinkGenedatabaseUniGeneTraceArchiveMapViewerHomologeneTheNewHomologeneAutomateddetectionofhomologsamongtheannotatedgenesofcompletelysequencedeukaryoticgenomes.NolongerUniGenebasedProteinsimilaritiesfirstGuidedbytaxonomictreeIncludesorthologsandparalogs
Orthologs和Paralogs是同源序列的兩種類型。
Orthologs(垂直同源基因)是指來自于不同物種的由垂直家系(物種形成)進化而來的蛋白,并且典型的保留與原始蛋白有相同的功能。Paralogs(平行同源基因)是那些在一定物種中的來源于基因復制的蛋白,可能會進化出新的與原來有關的功能。請參考文獻獲得更多的信息。geneduplicationParalogsvsOrthologsearlyglobingeneA-chaingene B-chaingenefrogAchickAmouseAmouse
BchickBfrogBparalogsorthologs
orthologsTheNewHomologene
HomologeneBuild37.2Species Numberofgenes inputgrouped groupsRAG1→Homologenerag112recombinationactivatinggeneRAG1→HomolgeneRAG1AmniotaHomolgene:RAG1Homolgene:RAG1GenomeResourcesLocusLinkGenedatabaseUniGeneTraceArchiveMapViewerHomologeneMapViewerListViewHumanMapVieweradar腺甙脫氨酶MapViewer:HumanADAR4MVHsADAR3’UTR5’UTRMaps&Options--Sequencemaps--AbinitioAssemblyRepeatsBES_CloneCloneNCI_CloneContigComponentCpGislanddbSNPhaplotypeFosmidGenBank_DNAGenePhenotypeSAGE_TagSTSTCAG_RNATranscript(RNA)Hs_UniGeneHs_EST--Cytogeneticmaps--IdeogramFISHCloneGene_CytogeneticMitelmanBreakpointMorbid/Disease--GeneticMaps--deCODEGenethonMarshfield--RHmaps--GeneMap99-G3GeneMap99-GB4NCBIRHStandford-G3TNGWhitehead-RHWhitehead-YACMm_UniGeneMm_ESTRn_UniGeneRn_ESTSsc_UniGeneSsc_ESTBt_UniGeneBt_ESTGga_UniGeneGga_ESTVariationMaps&Options=SNPMapViewerUniGeneComponentRepeatsGeneMastermap:repeatsGenePhenotypeVariationMaps&OptionsMaps&OptionsGenomeResourcesLocusLinkGenedatabaseUniGeneTraceArchiveMapViewerHomologeneStrongylocentrotuspurpuratus
TracesBLASTBasicLocalAlignmentSearchToolWebAccessBLASTVASTEntrezTextSequenceStructureBasicLocalAlignmentSearchToolWhyusesequencesimilarityBLASTalgorithmBLASTstatisticsBLASToutputExamplesWhyDoWeNeed
SequenceSimilaritySearchingToidentifyandannotatesequencesToevaluateevolutionaryrelationshipsOther:modelgenomicstructure(e.g.,Spidey)checkprimerspecificityinsilico:NCBI’stoolBLASTWebsiteStatsGlobalvsLocalAlignmentSeq1Seq2Seq1Seq2GlobalalignmentLocalalignmentGlobalvsLocalAlignmentSeq1:WHEREISWALTERNOW(16aa)Seq2:HEWASHEREBUTNOWISHERE(21aa)GlobalSeq1: 1W--HEREISWALTERNOW16 WHERESeq2: 1HEWASHEREBUTNOWISHERE21LocalSeq1:1W--HERE5Seq1:1W--HERE5WHEREWHERESeq2:3WASHERE9Seq2:15WISHERE21TheFlavorsofBLASTStandardBLASTtraditional“contiguous”wordhitpositionindependentscoringnucleotide,proteinandtranslations(blastn,blastp,blastx,tblastn,tblastx)MegablastoptimizedforlargebatchsearchescanusediscontiguouswordsPSI-BLASTconstructsPSSMsautomatically;usesasqueryverysensitiveproteinsearchRPSBLASTsearchesadatabaseofPSSMstoolforconserveddomainsearchesWidelyusedsimilaritysearchtoolHeuristicapproachbasedonSmithWatermanalgorithmFindsbestlocalalignmentsProvidesstatisticalsignificanceAllcombinations(DNA/Protein)queryanddatabase.DNAvsDNA
blastnDNAtranslationvsProtein
blastxProteinvsProtein
blastpProteinvsDNAtranslation
tblastnDNAtranslationvsDNAtranslation
tblastx
www,standalone,andnetworkclientsBasicLocalAlignmentSearchToolTranslatedBLASTQueryDatabaseProgramNPucleotideroteinNNNNPPblastxtblastntblastxPPPPPPPPPPPPPPPPPPPPPPPPParticularlyusefulfornucleotidesequenceswithoutproteinannotations,suchasESTsorgenomicDNAHowBLASTWorksMakelookuptableof“words”forqueryScandatabaseforhitsUngappedextensionsofhits(initialHSPs)Gappedextensions(notraceback)Gappedextensions(traceback;alignmentdetails)NucleotideWordsGTACTGGACATGGACCCTACAGGAAQuery:GTACTGGACATTACTGGACATGACTGGACATGGCTGGACATGGATGGACATGGACGGACATGGACCGACATGGACCCACATGGACCCTMakealookuptableofwords11-mer...828megablast711blastnminimumdefaultWORDSIZEProteinWordsGTQITVEDLFYNIATRRKALKNQuery:
NeighborhoodWordsLTV,MTV,ISV,LSV,etc.GTQTQIQITITVTVEVEDEDLDLF...MakealookuptableofwordsWordsize=3(default)Wordsizecanonlybe2or3[-f11=blastpdefault]MinimumRequirementsforaHit
NucleotideBLASTrequiresoneexactmatchProteinBLASTrequirestwoneighboringmatcheswithin40aaGTQITVEDLFYNI
SEIYYNATCGCCATGCTTAATTGGGCTT
CATGCTTAATT
neighborhoodwordsoneexactmatchtwomatches[-A40=blastpdefault]BLASTPSummary
YLS
HFLSbjct287LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEI333Query1IETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESI47GappedextensionwithtracebackQuery1IETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESI-LEV…50
+EYAYLKF+YLSL+SP++DVNVHP+KVHFL+++I++Sbjct287LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEIATSI…337FinalHSP
+EYAYLKF+
L+SP++DVNVHP+KV
+++I
High-scoringpair(HSP)HFL18HFV15HFS14HWL13NFL13DFL12HWV10etc…YLS15YLT12
YVS12YIT10etc…NeighborhoodwordsNeighborhoodscorethresholdT(-f)=11Query:
IETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESILEV…examplequerywordsScoringSystems-Nucleotides
AGCTA+1–3–3-3G–3+1–3-3C–3–3+1-3T–3–3–3+1IdentitymatrixCAGGTAGCAAGCTTGCATGTCA||||||||||||||||||| rawscore=19-9=10CACGTAGCAAGCTTG-GTGTCA[-r1-q-3]ScoringSystems-ProteinsPositionIndependentMatricesPAMMatrices(PercentAcceptedMutation)Derivedfromobservation;smalldatasetof alignmentsImplicitmodelofevolutionAllcalculatedfromPAM1PAM250widelyusedBLOSUMMatrices(BLOck
SUbstitution
Matrices)Derivedfromobservation;largedatasetofhighly conservedblocksEachmatrixderivedseparatelyfromblockswitha definedpercentidentitycutoffBLOSUM62-defaultmatrixforBLASTPositionSpecificScoreMatrices(PSSMs)
PSI-andRPS-BLASTA4R-15
N-206D-2-216C0-3-3-39Q-1100-35E-1002-425G0-20-1-3-2-26H-201-1-300-28I-1-3-3-3-1-3-3-4-34
L-1-2-3-4-1-2-3-4-324K-120-1-311-2-1-3-25M-1-1-2-3-10-2-3-212-15F-2-3-3-3-2-3-3-3-100-306P-1-2-2-1-3-1-1-2-2-3-3-1-2-47S1-110-1000-1-2-20-1-2-14T0-10-1-1-1-1-2-2-1-1-1-1-2-115W-3-3-4-4-2-2-3-2-2-3-2-3-11-4-3-211Y-2-2-2-3-2-1-2-32-1-1-2-13-3-2-227V0-3-3-3-1-2-2-3-331-21-1-2-20-3-14X0-1-1-1-2-1-1-1-1-1-1-1-1-1-200-2-1-1-1ARNDCQEGHILKMFPSTWYVXBLOSUM62DFNegativeforlesslikelysubstitutionsDYFPositiveformorelikelysubstitutionsPosition-SpecificScoreMatrixDAF-1Serine/Threonineproteinkinasescatalyticloop174PSSMscores54
ARNDCQEGHILKMFPSTWYV435K-100-1-23030-2-21-1-1-1-1-1-1-1-2436E0102-102-10-1-1000-100-1-1-1437S00-10110110-100020-1-10-1438N-10-1-110-133-1-11-100-1-111-1439K-211-1-20-1-2-2-1-251-2-2-1-1-2-2-1440P-2-2-2-2-3-2-2-2-2-1-2-10-37-1-2-3-1-1441A3-21-20-101-2-2-20-1-2310-3-30442M-3-4-4-4-3-4-4-5-470-410-4-4-2-4-12443A4-4-4-40-4-4-3-44-1-4-2-3-4-1-2-4-34
444H-4-2-1-3-5-2-2-410-6-5-3-4-3-2-3-4-50-5
445R-48-3-40-1-2-3-2-5-40-3-2-4-3-30-4-5
446D-4-4-18-6-20-3-3-5-6-3-5-6-4-2-3-7-5-5
447I-4-5-6-6-3-4-5-6-535-511-5-5-3-4-31
448K001-3-5-1-1-3-3-5-57-4-5-3-1-2-5-4-4
449S0-3-2-30-2-2-3-3-4-4-2-4-5262-5-4-4
450K0301-500-4-1-4-34-3-221-1-5-4-4
451N-4-38-1-5-2-2-3-1-6-6-2-4-5-4-1-2-6-4-5
452I-3-5-5-60-5-5-6-562-52-2-5-4-3-5-33
453M-4-4-6-6-3-4-5-6-506-510-5-4-3-4-30454V-3-3-5-6-3-4-5-6-533-42-2-5-4-3-5-35455K-2114-50-1-21-4-24-3-2-30-1-5-2-3456N1130-4-110-3-4-43-2-5-22-2-5-4-4457D-3-255-1-11-10-5-40-2-5-10-2-6-4-5458L-3-10-30-3-23-4-23011-2-2-35-1-3Position-SpecificScoreMatrixcatalyticloop[>./blastpgp-iNP_499868.2-dnr-j3-QNP_499868.pssm]LocalAlignmentStatisticsHighscoresoflocalalignmentsbetweentworandomsequencesfollowtheExtremeValueDistributionScore(S)Alignments(appliestoungappedalignments)E=Kmne-SorE=mn2-S’K=scaleforsearchspace=scaleforscoringsystemS’=bitscore=(S-lnK)/ln2ExpectValueE=numberofdatabasehitsyouexpecttofindbychance,≥SyourscoreexpectednumberofrandomhitsMoreinfo:
AdvancedBLASTOptions:NucleotideExampleEntrezQueriesnucleotideall[Filter]NOTmammalia[Organism]greenplants[Organism]biomolmrna[Properties]gbdivest[Properties]ANDrat[organism]OtherAdvanced–e10000 expectvalue-v2000 descriptions-b2000 alignmentsAdvancedBLASTOptions:ProteinMatrixSelectionPAM30--moststringentBLOSUM45--leaststringentExampleEntrezQueriesproteinsall[Filter]NOTmammalia[Organism]greenplants[Organism]srcdbrefseq[Properties]OtherAdvanced–e10000 expectvalue-v2000 descriptions-b2000 alignmentsLimitbytaxonMusmusculus[Organism]Mammalia[Organism]Viridiplantae[Organism]sp|P27476|NSR1_YEASTNUCLEARLOCALIZATIONSEQUENCEBINDINGPROTEIN(P67)Length=414Score=40.2bits(92),Expect=0.013Identities=35/131(26%),Positives=56/131(42%),Gaps=4/131(3%)Query:362STTSLTSSSTSGSSDKVYAHQMVRTDSREQKLDAFLQPLSKPLSSQPQAIVTEDKTD418S++SSSS+SSS++++S++SSS++EKSbjct:29SSSSSESSSSSSSSSESESESESESESSSSSSSSDSESSSSSSSDSESEAETKKEESKDS88FilteredUnfilteredLowComplexityFilteringOtherBLASTAlgorithms
MegablastDiscontiguousMegablastPSI-BLASTPHI-BLASTMegablast:NCBI’sGenomeAnnotator
LongalignmentsofsimilarDNAsequencesGreedyalgorithmConcatenationofquerysequencesFasterthanblastn;lesssensitiveMegaBLAST&WordSizeTrade-off:sensitivityvsspeed23blastp828megablast711blastnminimumdefaultWORDSIZEDiscontiguousMegablastUsesdiscontiguouswordmatchesBetterforcross-speciescomparisonsTemplatesforDiscontiguousWordsW=11,t=16,coding: 1101101101101101W=11,t=16,non-coding: 1110010110110111W=12,t=16,coding: 1111101101101101W=12,t=16,non-coding: 1110110110110111W=11,t=18,coding: 101101100101101101W=11,t=18,non-coding: 111010010110010111W=12,t=18,coding: 101101101101101101W=12,t=18,non-coding: 111010110010110111W=11,t=21,coding: 100101100101100101101W=11,t=21,non-coding: 111010010100010010111W=12,t=21,coding: 100101101101100101101W=12,t=21,non-coding: 111010010110010010111Reference:Ma,B,Tromp,J,Li,M.PatternHunter:fasterandmoresensitivehomologysearch.BioinformaticsMarch,2002;18(3):440-5W=wordsize;#matchesintemplatet=templatelengthDiscontiguous(Cross-species)MegaBLASTDiscontiguousWordOptionsMegaBLASTvsDiscontiguousMegaBLASTNM_017460HomosapienscytochromeP450,family3,subfamilyA,polypeptide4(CYP3A4),transcriptvariant1,mRNA(2768letters)vsDrosophilaMegaBLASTvsDiscontiguousMegaBLAST
MegaBLAST=“Nosignificantsimilarityfound.”
Discontiguous
megaBLAST=AnotherExample...
Discontiguous
megaBLAST=numeroushits...Query:NM_078651DrosophilamelanogasterCG18582-PA(mbt)mRNA,(3244bp)/note=mushroombodiestiny;synonyms:Pak2,STE20,dPAK2
MegaBLAST=“Nosignificantsimilarityfound.”Database:nr(nt),Mammalia[orgn]Ex:DiscontiguousMegaBLASTEx:BLASTNPSI-BLASTExample:ConfirmingrelationshipsofpurinenucleotidemetabolismproteinsPosition-specificIteratedBLAST>gi|113340|sp|P03958|ADA_MOUSEADENOSINEDEAMINASE(ADENOSINEMAQTPAFNKPKVELHVHLDGAIKPETILYFGKKRGIALPADTVEELRNIIGMDKPLSLPGFVIAGCREAIKRIAYEFVEMKAKEGVVYVEVRYSPHLLANSKVDPMPWNQTEGDVTPDDVVDEQAFGIKVRSILCCMRHQPSWSLEVLELCKKYNQKTVVAMDLAGDETIEGSSLFPGHVEAYRTVHAGEVGSPEVVREAVDILKTERVGHGYHTIEDEALYNRLLKENMHFEVCPWSSYLTGAVRFKNDKANYSLNTDDPLIFKSTLDTDYQMTKKDMGFTEEEFKRLNINAAKSSFLPEEEKKPSI-BLAST0.005EvaluecutoffforPSSMRESULTS:InitialBLASTPSameresultsasprotein-proteinBLAST;differentformatResultsofFirstPSSMSearchOtherpurinenucleotidemetabolizingenzymesnotfoundbyordinaryBLASTTenthPSSMSearch:ConvergenceJustbelowthreshold,anothernucleotidemetabolismenzymeChecktoaddtoPSSMReversePSI-BLAST(RPS)-BLASTAdenosine/AMPDeaminaseDomainAMPDeaminases...PHI-BLAST>gi|231729|sp|P30429|CED4_CAEELCELLDEATHPROTEIN4MLCEIECRALSTAHTRLIHDFEPRDALTYLEGKNIFTEDHSELISKMSTRLERIANFLRIYRRQASELIDFFNYNNQSHLADFLEDYIDFAINEPDLLRPVVIAPQFSRQMLDRKLLLGNVPKQMTCYIREYHVIKKLDEMCDLDSFFLFLHGRAGSGKSVIASQALSKSDQLIGINYDSIVWLKDSGTAPKSTFDLFTDILKSEDDLLNFPSVEHVTSVVLKRMICNALIDRPNTLFVFDDVVQEETIRWAQELRLRCLVTTRDVEIASQTCEFIEVTSLEIDECYDFLEAYGMPMPVGEKEEDVLNKTIELSSGNPATLMMFFKSCEPKTFEK[GA]xxxxGK[ST]What’sNewBLASTDatabasesNucleotiderefseq_rna=NM_*,XM_*refseq_genomic=NC_*,NG_*env_ntenvironmentalsample[filter],e.g.,16SrRNAProteinrefseq=NP_*,XP_*env_nrNewFormatterSelectlowercaseSelectredNewFormattergrayline=samedatabasehit
hsp’scolor-codedindependentlyBLASTOutput:Alignments&FilterlowcomplexitysequencefilteredAdvanc
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