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An Introduction to An Introduction to Metagenomics Data AnalysisMetagenomics Data Analysis
Metagenomics TrainingMetagenomics Training
Ferran BriansóFerran Briansó
VHIR - 26/08/2013
[email protected]@vhir.org
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OutlineOutline
Introduction to Metagenomics
Basic Terminology
Computational Approaches & Tools Whole Genome Shotgun 16S/ITS Community Surveys
Recommended Tools MEGAN mothur QIIME AXIOME & CloVR
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Introduction to METAGENOMICSMETAGENOMICS
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IntroductionIntroduction
First use of the term metagenome, referencing the idea that a collection of genes sequenced from the environment could be analyzed in a way analogous to the study of a single genome.
Handelsman, J.; Rondon, M. R.; Brady, S. F.; Clardy, J.; Goodman, R. M. (1998). "Molecular biological access to the chemistry of unknown soil microbes: A new frontier for natural products".Chemistry & Biology 5 (10): R245–R249. doi:10.1016/S1074-5521(98)90108-9. PMID 9818143
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First use of the term metagenome, referencing the idea that a collection of genes sequenced from the environment could be analyzed in a way analogous to the study of a single genome.
“The application of modern genomics techniques to the study of communities of microbial organisms directly in their natural environments, bypassing the need for isolation and lab cultivation of individual species.”
Handelsman, J.; Rondon, M. R.; Brady, S. F.; Clardy, J.; Goodman, R. M. (1998). "Molecular biological access to the chemistry of unknown soil microbes: A new frontier for natural products".Chemistry & Biology 5 (10): R245–R249. doi:10.1016/S1074-5521(98)90108-9. PMID 9818143
Chen, K.; Pachter, L. (2005). "Bioinformatics for Whole-Genome Shotgun Sequencing of Microbial Communities".PLoS Computational Biology 1 (2): e24. doi:10.1371/journal.pcbi.0010024
IntroductionIntroduction
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Source: US Division of Earth & Life Studies of the National Academieshttp://dels-old.nas.edu/metagenomics/overview.shtml
IntroductionIntroduction
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Source: US Division of Earth & Life Studies of the National Academieshttp://dels-old.nas.edu/metagenomics/overview.shtml
IntroductionIntroduction
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Source:
IntroductionIntroduction
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Source: Feng Chen, JGI
IntroductionIntroduction
Perfomance Comparison for (some) Platforms
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Basic TERMINOLOGYTERMINOLOGY
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TerminologyTerminology
Trimming: is the pre-processing step of cleaning sequence data (primers, multiplexing barcodes...) from automated DNA sequencers prior to sequence assembly and other downstream uses.
Binning is the process of grouping reads or contigs and assigning them to operational taxonomic units (OTUs).
OTU (Operational Taxonomic Unit): Taxonomic level of sampling selected by the user to be used in a study. Typically using a percent sequence similarity threshold for classifying microbes within the same, or different, OTUs.
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Trimming: is the pre-processing step of cleaning sequence data (primers, multiplexing barcodes...) from automated DNA sequencers prior to sequence assembly and other downstream uses.
Binning is the process of grouping reads or contigs and assigning them to operational taxonomic units (OTUs).
OTU (Operational Taxonomic Unit): Taxonomic level of sampling selected by the user to be used in a study. Typically using a percent sequence similarity threshold for classifying microbes within the same, or different, OTUs.
Chimeras: Artificial sequences formed during PCR amplification. The majority of them are believed to arise from incomplete extension. During subsequent cycles of PCR, a partially extended strand can bind to a template derived from a different but similar sequence. This then acts as a primer that is extended to form a chimeric sequence (Smith et al. 2010, Thompson et al., 2002, Meyerhans et al., 1990, Judo et al., 1998, Odelberg, 1995). A chimeric template is created during one round, then amplified by subsequent rounds to produce chimeric amplicons that are difficult to distinguish from amplicons derived from a single biological sequence.
TerminologyTerminology
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Trimming: is the pre-processing step of cleaning sequence data (primers, multiplexing barcodes...) from automated DNA sequencers prior to sequence assembly and other downstream uses.
Binning is the process of grouping reads or contigs and assigning them to operational taxonomic units (OTUs).
OTU (Operational Taxonomic Unit): Taxonomic level of sampling selected by the user to be used in a study. Typically using a percent sequence similarity threshold for classifying microbes within the same, or different, OTUs.
Chimeras: Artificial sequences formed during PCR amplification. The majority of them are believed to arise from incomplete extension. During subsequent cycles of PCR, a partially extended strand can bind to a template derived from a different but similar sequence. This then acts as a primer that is extended to form a chimeric sequence (Smith et al. 2010, Thompson et al., 2002, Meyerhans et al., 1990, Judo et al., 1998, Odelberg, 1995). A chimeric template is created during one round, then amplified by subsequent rounds to produce chimeric amplicons that are difficult to distinguish from amplicons derived from a single biological sequence.
Alpha diversity: the diversity within a particular area or ecosystem; expressed by the number of species (i.e., species richness) in that ecosystem, or by one or more diversity indices.
Beta diversity: a comparison of of diversity between ecosystems, usually measured as the amount of species change between the ecosystems.
Gamma diversity: a measure of the overall diversity within a large region. Geographic-scale species diversity according to Hunter (2002:448).
TerminologyTerminology
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Trimming: is the pre-processing step of cleaning sequence data (primers, multiplexing barcodes...) from automated DNA sequencers prior to sequence assembly and other downstream uses.
Binning is the process of grouping reads or contigs and assigning them to operational taxonomic units (OTUs).
OTU (Operational Taxonomic Unit): Taxonomic level of sampling selected by the user to be used in a study. Typically using a percent sequence similarity threshold for classifying microbes within the same, or different, OTUs.
Chimeras: Artificial sequences formed during PCR amplification. The majority of them are believed to arise from incomplete extension. During subsequent cycles of PCR, a partially extended strand can bind to a template derived from a different but similar sequence. This then acts as a primer that is extended to form a chimeric sequence (Smith et al. 2010, Thompson et al., 2002, Meyerhans et al., 1990, Judo et al., 1998, Odelberg, 1995). A chimeric template is created during one round, then amplified by subsequent rounds to produce chimeric amplicons that are difficult to distinguish from amplicons derived from a single biological sequence.
Alpha diversity: the diversity within a particular area or ecosystem; expressed by the number of species (i.e., species richness) in that ecosystem, or by one or more diversity indices.
Beta diversity: a comparison of of diversity between ecosystems, usually measured as the amount of species change between the ecosystems.
Gamma diversity: a measure of the overall diversity within a large region. Geographic-scale species diversity according to Hunter (2002:448).
Rarefaction allows the calculation of species richness for a given number of individual samples, based on the construction of so-called rarefaction curves. This curve is a plot of the number of species as a function of the number of samples.
TerminologyTerminology
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Computational APPROACHES & TOOLSAPPROACHES & TOOLS
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Approaches & ToolsApproaches & Tools
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Approaches & ToolsApproaches & Tools
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Approaches & ToolsApproaches & Tools
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Approaches & ToolsApproaches & Tools
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Whole Genome SHOTGUNSHOTGUN
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Whole Genome ShotgunWhole Genome Shotgun
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WGS WorkflowWGS Workflow
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WGS WorkflowWGS Workflow
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WGS WorkflowWGS Workflow
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WGS WorkflowWGS Workflow
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Examples of WGS ToolsExamples of WGS Tools
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Examples of WGS ToolsExamples of WGS Tools
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Analysis of 16S/ITS 16S/ITS Community SurveysCommunity Surveys
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16S/ITS community surveys16S/ITS community surveys
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16S/ITS issues16S/ITS issues
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16S/ITS workflow16S/ITS workflow
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16S/ITS workflow16S/ITS workflow
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16S/ITS workflow16S/ITS workflow
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16S/ITS workflow16S/ITS workflow
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Some recommended ToolsTools
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Some (recommended) ToolsSome (recommended) Tools
mothur
MEGAN
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MEGANMEGAN
2007 →
2011 →
...
...
2012 →
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MEGAN 4 for 16S rRNAMEGAN 4 for 16S rRNA
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MEGAN 4 for 16S rRNAMEGAN 4 for 16S rRNA
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mothurmothur
2009 →
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mothurmothur
2009 →
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QIIMEQIIME
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Integrative Tools/PlatformsTools/Platforms
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AXIOMEAXIOME
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AXIOMEAXIOME
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AXIOMEAXIOME
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CloVRCloVR
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CloVRCloVR
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CloVRCloVR
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CloVRCloVR
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CloVRCloVR
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Ferran BriansóFerran BriansóMGTraining 26/08/2013
Thanks for your attentionThanks for your attention
[email protected]@vhir.org
An Introduction to An Introduction to Metagenomics Data AnalysisMetagenomics Data Analysis
more info at http://ueb.vhir.org/MGT