The effects of alignment quality, distance calculation method, sequence filtering, and region on the analysis of 16S rRNA gene-based studies.

PLoS Computational Biology
Patrick D Schloss

Abstract

Pyrosequencing of PCR-amplified fragments that target variable regions within the 16S rRNA gene has quickly become a powerful method for analyzing the membership and structure of microbial communities. This approach has revealed and introduced questions that were not fully appreciated by those carrying out traditional Sanger sequencing-based methods. These include the effects of alignment quality, the best method of calculating pairwise genetic distances for 16S rRNA genes, whether it is appropriate to filter variable regions, and how the choice of variable region relates to the genetic diversity observed in full-length sequences. I used a diverse collection of 13,501 high-quality full-length sequences to assess each of these questions. First, alignment quality had a significant impact on distance values and downstream analyses. Specifically, the greengenes alignment, which does a poor job of aligning variable regions, predicted higher genetic diversity, richness, and phylogenetic diversity than the SILVA and RDP-based alignments. Second, the effect of different gap treatments in determining pairwise genetic distances was strongly affected by the variation in sequence length for a region; however, the effect of different calcul...Continue Reading

References

Mar 1, 1970·Journal of Molecular Biology·S B Needleman, C D Wunsch
Dec 10, 2002·Proceedings of the National Academy of Sciences of the United States of America·R A WelchF R Blattner
Dec 9, 2003·Molecular Biology and Evolution·Andrew D SmithElisabeth R M Tillier
Mar 23, 2004·Nucleic Acids Research·Robert C Edgar
Apr 16, 2005·Science·Paul B EckburgDavid A Relman
Aug 2, 2005·Nature·Marcel MarguliesJonathan M Rothberg
Jul 6, 2006·Applied and Environmental Microbiology·T Z DeSantisG L Andersen
Jul 18, 2006·Nucleic Acids Research·T Z DeSantisG L Andersen
Jul 20, 2006·PLoS Computational Biology·Patrick D Schloss, Jo Handelsman
Aug 2, 2006·Proceedings of the National Academy of Sciences of the United States of America·Mitchell L SoginGerhard J Herndl
Sep 20, 2006·Bioinformatics·Luke ShenemanJames A Foster
Jun 26, 2007·Applied and Environmental Microbiology·Qiong WangJames R Cole
Sep 21, 2007·Nucleic Acids Research·Zongzhi LiuRob Knight
Oct 6, 2007·Science·Julie A HuberMitchell L Sogin
Nov 29, 2007·The ISME Journal·Luiz F W RoeschEric W Triplett
Nov 14, 2008·Nucleic Acids Research·J R ColeJ M Tiedje
May 7, 2009·Nucleic Acids Research·Yijun SunWilliam Farmerie
Nov 7, 2009·Science·Elizabeth K CostelloRob Knight
Nov 17, 2009·Bioinformatics·J Gregory CaporasoRob Knight

❮ Previous
Next ❯

Citations

Apr 26, 2014·PLoS Computational Biology·Thomas S B SchmidtChristian von Mering
Sep 14, 2013·Journal of Microbiological Methods·Julia M Di BellaGregor Reid
Sep 2, 2010·Microbiology and Molecular Biology Reviews : MMBR·Courtney J RobinsonVincent B Young
Mar 23, 2011·Applied and Environmental Microbiology·Patrick D Schloss, Sarah L Westcott
Sep 6, 2011·Applied and Environmental Microbiology·David BerryAlexander Loy
May 24, 2012·Nucleic Acids Research·Bernd W BrandtEgija Zaura
Sep 29, 2012·The ISME Journal·Patrick D Schloss
Dec 5, 2012·PLoS Computational Biology·Dirk GeversCurtis Huttenhower
Jan 10, 2013·PLoS Computational Biology·Xochitl C Morgan, Curtis Huttenhower
Dec 13, 2012·Microbial Ecology·Nastassia V PatinMatthew N Ashby
Jul 13, 2012·Briefings in Bioinformatics·Monica SantamariaGraziano Pesole
Sep 15, 2012·The ISME Journal·Viggó Thór MarteinssonEric Gaidos
Dec 3, 2013·Indian Journal of Microbiology·Balakrishnan SundarakrishnanParamasamy Gunasekaran
Sep 16, 2015·Microbial Ecology·Yu-Ling HuangA Elizabeth Arnold
Jan 12, 2016·Scientific Reports·Anna Maria Fiore-DonnoMichael Bonkowski
Nov 5, 2010·Journal of Microbiological Methods·Minseok KimZhongtang Yu
Sep 1, 2015·Bioinformatics·Rujira AchawanantakunYuan Zhang
Nov 29, 2015·Current Microbiology·Arumugam ManjulaJeyaprakash Rajendhran
Feb 22, 2012·Neurogastroenterology and Motility : the Official Journal of the European Gastrointestinal Motility Society·I M CarrollY Ringel

❮ Previous
Next ❯

Methods Mentioned

BETA
PCR

Software Mentioned

clearcut
DNADIST
mothur
MUSCLE
kmer
SILVA
UniFrac
greengenes
PHYLIP

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.