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EMIRGE reconstruction of full-length ribosomal genes from microbial community short read sequencing data.doc

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EMIRGE reconstruction of full-length ribosomal genes from microbial community short read sequencing data

Milleretal.GenomeBiology2011,12:R44 /2011/12/5/R44 METHOD OpenAccess EMIRGE:reconstructionoffull-lengthribosomal genesfrommicrobialcommunityshortread sequencingdata ChristopherSMiller1*,BrettJBaker1,2,BrianCThomas1,StevenWSinger3,4 andJillianFBanfield1,5* Abstract RecoveryofribosomalsmallsubunitgenesbyassemblyofshortreadcommunityDNAsequencedatagenerally fails,makingtaxonomiccharacterizationdifficult.Here,wesolvethisproblemwithanoveliterativemethod,based ontheexpectationmaximizationalgorithm,thatreconstructsfull-lengthsmallsubunitgenesequencesand providesestimatesofrelativetaxonabundances.Weapplythemethodtonaturalandsimulatedmicrobial communities,andcorrectlyrecovercommunitystructurefromknownandpreviouslyunreportedrRNAgene sequences.Animplementationofthemethodisfreelyavailableat/csmiller/EMIRGE. Background not recovered by methods that relied on amplification Characterizationofmicrobialcommunitycompositionis withstandardSSUprimers. mostoftendonewithaphylogeneticmarkergene,most MostreportedmetagenomicsequencinghasusedSan- commonly the small subunit ribosomal RNA (SSU ger orRoche 454 sequencing technologies. The rRNA rRNA) gene [1]. Traditionally, rRNA sequences were genesequencesforcloselyrelatedorganismsinthesedata- generatedbyamplification,cloning,andSangersequen- setsco-assemble.Theresultisacompositesequencethat cing. More recently, technologies such as pyrotag is not representative of any community member and sequencingofshorthyper-variableregions[2,3],Illumina obscures thereallevel ofdiversity. Theseproblems are sequencing ofvariable tags [4-7], andhybridization to exacerbated when shorter sequencing reads are used. specializedhigh-densitymicroarrays(forexample,Phylo- TypicalreadsfromtheIlluminaplatformcurrentlyrange chip) [8-10] have accelerated the throughput of SSU- from35to125bp.Additionally,thek-mer-basedmethods basedmicrobial community characterization. Although usedtoassembleshortreaddatacanfurtherconfoundde each method has