Evaluation of Methods for the Extraction and Purification of DNA from the Human Microbiome 英文参考文献.docVIP
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Evaluation of Methods for the Extraction and Purification of DNA from the Human Microbiome 英文参考文献
EvaluationofMethodsfortheExtractionandPurification
ofDNAfromtheHumanMicrobiome
SanqingYuan1,4,DoraB.Cohen1,4,JacquesRavel5,ZaidAbdo2,3,4,LarryJ.Forney1,4
*
1DepartmentofBiologicalSciences,UniversityofIdaho,Moscow,Idaho,UnitedStatesofAmerica,2DepartmentofMathematics,UniversityofIdaho,Moscow,Idaho,
UnitedStatesofAmerica,3DepartmentofStatistics,UniversityofIdaho,Moscow,Idaho,UnitedStatesofAmerica,4InstituteforBioinformaticsandEvolutionaryStudies,
UniversityofIdaho,Moscow,Idaho,UnitedStatesofAmerica,5InstituteforGenomeSciences,UniversityofMarylandSchoolofMedicine,Baltimore,Maryland,United
StatesofAmerica
Abstract
Background: DNA extraction is an essential step in all cultivation-independent approaches to characterize microbial
diversity,includingthatassociatedwiththehumanbody.Afundamentalchallengeinusingtheseapproacheshasbeento
isolateDNAthatisrepresentativeofthemicrobialcommunitysampled.
Methodology/Principal Findings:Inthisstudy,westatisticallyevaluatedsixcommonlyusedDNAextractionprocedures
using elevenhuman-associated bacterial species andamockcommunity that containedequal numbers ofthose eleven
species. Thesemethods were compared on the basis of DNA yield, DNA shearing, reproducibility, and most importantly
representationofmicrobialdiversity.Theanalysisof16SrRNAgenesequencesfromamockcommunityshowedthatthe
observedspeciesabundancesweresignificantlydifferentfromtheexpectedspeciesabundancesforallsixDNAextraction
methodsused.
Conclusions/Significance:Protocolsthatincludedbeadbeatingand/ormutanolysinproducedsignificantlybetterbacterial
communitystructurerepresentationthanmethodswithoutbothofthem.Thereproducibilityofallsixmethodswassimilar,
andresultsfromdifferentexperimenters anddifferenttimeswereingoodagreement.Basedontheevaluationsdoneit
appearsthatDNAextractionproceduresforbacterialcommunityanalysisofhumanassociatedsamplesshouldincludebead
beatingand/ormutanolysintoeffectivelylysecells.
Citation: Yuan S, Cohen DB, Ravel J, Abdo Z, Forney LJ (2012) Evaluation of Me
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