Genboree Microbiome Toolsetand Virtual Data Integration

Kevin P. RiehleMatthew E. Roth

Aleksandar MilosavljevicBaylor College of Medicine

April 2-4, 2012Boulder, Colorado

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Genboree Introduction

• Genboree.org– Everyone should have received an email regarding

their Genboree account• If not, ask or email us (riehle@bcm.edu)

• Genboree.org/microbiome– Tutorial– NIH Cloud Workshop Material– FAQ

• Links also provided on the shared Google docAleksandar Milosavljevic, Baylor College of

Medicine amilosav@bcm.edu

Invited to BCM Bioinformatics:

Additional contributors not acknowledged in the reference above:

R. Alan Harris, Tim Charnecki

Funding:

This project was supported in part by the NIH Roadmap Epigenomics (Common Fund) U01 DA025956 and NIH-NHGRI R01HG004009 grants to AM, by the NIH-NIDDK UH3 DK083990, P30 DK56338, and NIH Common Fund and NIH-NHGRI U54 HG003273 and U54 HG004973 grants to JV, and by Burroughs Welcome Fund Preterm Birth Initiative and NIH DP21DP2OD001500-01 grants to KA..

References and Acknowledgments

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Login at www.genboree.org

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Access Microbiome Toolset

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Genboree SNP / Resequencing Toolset

BMC Genomics (Invited submission)

Publications

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Genboree RNA Toolset

Publications enabled by the Genboree small RNA Toolset• Gunaratne PH, Lin YC, Benham AL, Drnevich J, Coarfa C, Tennakoon JB, Creighton CJ, Kim JH, Milosavljevic A, Watson M, Griffiths-Jones

S, Clayton DF. “Song exposure regulates known and novel microRNAs in the zebra finch auditory forebrain.” BMC Genomics. 2011 May 31;12(1):277.

• Shohet JM, Ghosh R, Coarfa C, Ludwig A, Benham AL, Chen Z, Patterson DM, Barbieri E, Mestdagh P, Sikorski DN, Milosavljevic A, Kim ES, Gunaratne PH. “A genome-wide search for promoters that respond to increased MYCN reveals both new oncogenic and tumor suppressor microRNAs associated with aggressive neuroblastoma. “ Cancer Res. 2011 Jun 1;71(11):3841-51.

• Buchold GM, Coarfa C, Kim J, Milosavljevic A, Gunaratne PH, Matzuk MM. “Analysis of microRNA expression in the prepubertal testis. “ PLoS One. 2010 Dec 29;5(12):e15317.

• Polikepahad S, Knight JM, Naghavi AO, Oplt T, Creighton CJ, Shaw C, Benham AL, Kim J, Soibam B, Harris RA, Coarfa C, Zariff A, Milosavljevic A, Batts LM, Kheradmand F, Gunaratne PH, Corry DB. “Proinflammatory role for let-7 microRNAS in experimental asthma.” J Biol Chem. 2010 Sep 24;285(39):30139-49.

• Ma L, Buchold GM, Greenbaum MP, Roy A, Burns KH, Zhu H, Han DY, Harris RA, Coarfa C, Gunaratne PH, Yan W, Matzuk MM. “GASZ is essential for male meiosis and suppression of retrotransposon expression in the male germline.” PLoS Genet. 2009 Sep;5(9):e1000635. Epub 2009 Sep 4. Erratum in: PLoS Genet. 2009 Dec;5(12).

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Genboree Epigenomics Toolset

Human Epigenome Atlas and the Genboree Epigenomic Toolset for Comparative Epigenome AnalysisCoarfa C1, Harris RA1, Jackson AR1, Pichot CS2, Raghuraman S1, Paithankar S1, Lee AV3, McGuire SE2, Milosavljevic A1

1NIH Roadmap Epigenomics Data Analysis and Coordination Center (EDACC), Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas. 2Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas. 3Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, Pennsylvania..

The NIH Roadmap Epigenomics Program Investigators’ Meeting, May 14-15 2012, Bethesda, Maryland, USA.

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Hosts / Domains

Groups Databases Annotations Sample sets Files

Projects

Launch Genboree Workbench

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Hosts / Domains

Groups Databases Annotations Sample sets Files

Projects

Managing, Groups, Projects and DatabasesCreateReadUpdateDelete

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Hosts / Domains

Groups Databases Annotations Sample sets Files

Projects

Launch Genboree Workbench

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

Virtual Data Integration

www.microbiome-center.org

www.brain-research-lab.org

www.genboree.org

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Genboree Workbenches

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

Virtual Data Integration

www.microbiome-center.org

www.brain-research-lab.org

www.genboree.org

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Physical site #1 Physical site #2Physical site #3

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Virtual Data Integration via Genboree REST APIs

Compute Cluster

Databases & File Storage

Web Site

Servers

Genboree

REST API Server

Genboree Workbench UIs

(running in user’s browser)

REST API Executing Tool Job, Data Storage, Etc

Person 2

Person 2

wwww.genboree.org hosted at Baylor

www.brain-research-lab.org andwww.microbiome-center.org at Rackspace

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Virtual Data Integration

A combination of dedicated hosting and elastic cloud computing accessible via the Workbench

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Supplying Credentials to www.microbiome-center.org

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Virtual Integration

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Virtual Integration

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Virtual Integration

Drag and drop

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Virtual Integration

Drag and drop

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Data Tree Selector

Various Data Types

Item Details

Data Type Filter

Input Data

Output Targets

Genboree Workbench

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Activated Tool

Non-Activated Tool

Genboree Workbench

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16S Microbiome Toolset Work Flow

2. Transfer 3. Link Samples to Seq Files

5. Run Tools:• RDP• QIIME• Alpha

Diversity• Machine

Learning

4. Import Sequences

SRR, SFF Sequences

Sample Meta Data

Sample Record

Sample Record

Sample Record

Quality Filtered Sequences

1. Setup

Create:

• Sample Meta Data

• Group• Database• Project

α β

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16S Microbiome Toolset Tutorial1. Setup

• Create Sample Meta Data• Create Group• Create Database• Create Project

2. Transfer• Upload Files• View Uploaded Files• Import Samples• View Imported Samples

3. Link Samples to Sequence Files4. Import Sequences

• View Imported Sequences• Analysis

5. Run Tools• RDP – Taxonomic Abundance Pipeline• QIIME Pipeline – OTU Table, Phylogenetic Tree, and Beta Diversity• Alpha Diversity• Machine Learning

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Multi-step OTU Picking

Remove Chimeras

Taxonomic Classification

Representative Sequences

OTU Table Phylogenetic Tree

Beta Diversity

Alpha Diversity

Classification

Feature Selection

Taxonomic Abundance

= end results

= intermediate resultQuality Filtered Sequences

16S Microbiome

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Microbiome Tutorial1. Introduction to 16S Analysis

• Genboree.org/microbiome– http://genboree.org/theCommons/projects/pub-gen-microbiome/wiki/Microbiome_Tutorial

» 5 Stool samples» 5 Throat samples

2. Replicating Published Data• A core gut microbiome in obese and lean twins (Turnbaugh et al, Nature, 2009)• Study objective: To address the question of how host genotype,

environmental exposures, and host adiposity influence the gut microbiome – http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2677729/pdf/nihms74182.pdf

» 49 Lean Stool samples» 45 Obese Stool samples

– Comparing the alpha diversity between lean and obese stool samples

3. Utilizing HMP control data– > 13,000 samples– 18 body sites– 3 primer regions

4. KEGG Mashups– Subset of HMP WGS HUMAnN output

» 50 samples» 7 body sites

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Microbiome Tutorial1. Introduction to 16S Analysis

• Genboree.org/microbiome– http://genboree.org/theCommons/projects/pub-gen-microbiome/wiki/Microbiome_Tutorial

» 5 Stool samples» 5 Throat samples

2. Replicating Published Data• A core gut microbiome in obese and lean twins (Turnbaugh et al, Nature, 2009)• Study objective: To address the question of how host genotype,

environmental exposures, and host adiposity influence the gut microbiome – http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2677729/pdf/nihms74182.pdf

» 49 Lean Stool samples» 45 Obese Stool samples

– Comparing the alpha diversity between lean and obese stool samples

3. Utilizing HMP control data– > 13,000 samples– 18 body sites– 3 primer regions

4. KEGG Mashups– Subset of HMP WGS HUMAnN output

» 50 samples» 7 body sites

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Introduction to 16S Analysis

• http://www.Genboree.org/microbiome– http://genboree.org/theCommons/projects/pu

b-gen-microbiome/wiki/Microbiome_Tutorial• 5 Stool samples• 5 Throat samples

– Group• NIH_CLOUD_WORKSHOP

– Database• nih_workshop_tutorial

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Microbiome Tutorial1. Introduction to 16S Analysis

• Genboree.org/microbiome– http://genboree.org/theCommons/projects/pub-gen-microbiome/wiki/Microbiome_Tutorial

» 5 Stool samples» 5 Throat samples

2. Replicating Published Data• A core gut microbiome in obese and lean twins (Turnbaugh et al, Nature, 2009)• Study objective: To address the question of how host genotype,

environmental exposures, and host adiposity influence the gut microbiome – http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2677729/pdf/nihms74182.pdf

» 49 Lean Stool samples» 45 Obese Stool samples

– Comparing the alpha diversity between lean and obese stool samples

3. Utilizing HMP control data– > 13,000 samples– 18 body sites– 3 primer regions

4. KEGG Mashups– Subset of HMP WGS HUMAnN output

» 50 samples» 7 body sites

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Replicating Published Data

• Lean vs obese twin study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

• 94 samples– 49 Lean– 45 Obese

• V6 primer region• 454 – 16S rRNA

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Genboree Project Integration

• http://genboree.org/java-bin/project.jsp?projectName=Turnbaugh_lean_obese_twins_project

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Phylogenetic Visualizations - iTOL

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

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Lean vs. Obese Twins Study

Vol 457|22 January 2009| doi:10.1038/nature07540

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Lean vs. Obese Twins Study

Vol 457|22 January 2009| doi:10.1038/nature07540

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Microbiome Tutorial1. Introduction to 16S Analysis

• Genboree.org/microbiome– http://genboree.org/theCommons/projects/pub-gen-microbiome/wiki/Microbiome_Tutorial

» 5 Stool samples» 5 Throat samples

2. Replicating Published Data• A core gut microbiome in obese and lean twins (Turnbaugh et al, Nature, 2009)• Study objective: To address the question of how host genotype,

environmental exposures, and host adiposity influence the gut microbiome – http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2677729/pdf/nihms74182.pdf

» 49 Lean Stool samples» 45 Obese Stool samples

– Comparing the alpha diversity between lean and obese stool samples

3. Utilizing HMP control data– > 13,000 samples– 18 body sites– 3 primer regions

4. KEGG Mashups– Subset of HMP WGS HUMAnN output

» 50 samples» 7 body sites

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Grid Viewer

• Provides an interactive view of Samples from 1 to many databases

• Databases may exist in different physical locations (virtual integration) (will discuss more later)

• Users can save Sample Sets in which to analyze • Users can select Samples in which to explore

Genes and Pathways (WGS only)

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HMP Data Metrics

• Phase I and Phase II– http://trace.ncbi.nlm.nih.gov/Traces/sra/?

study=SRP002395– http://trace.ncbi.nlm.nih.gov/Traces/sra/?study=S

RP002860– > 13,000 samples

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Grid Viewer

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16S rRNA Sample Grid Viewer

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16S rRNA Sample Grid Viewer

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16S rRNA Sample Grid Viewer

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16S rRNA Sample Grid Viewer

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16S rRNA Sample Grid Viewer

• Then show how we can use these sample sets for analysis on the GMT

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16S rRNA Sample Grid Viewer• http://genboree.org/java-bin/sampleGridViewer.jsp?dbList=http%3A%2F

%2Fgenboree.org%2FREST%2Fv1%2Fgrp%2FHMP-16S-rRNA-phaseI-phaseII%2Fdb%2FHMP-16S-I-II%3F&gbGridXAttr=primer_region&gbGridYAttr=body_site&xlabel=primer_region&ylabel=body_site&gridTitle=Samples%20from%20HMP-16S-I-II&pageTitle=Sample%20Grid%20Viewer:%20Samples%20from%20HMP-16S-I-II

• http://genboree.org/java-bin/sampleGridViewer.jsp?dbList=http%3A%2F%2Fgenboree.org%2FREST%2Fv1%2Fgrp%2FHMP-16S-rRNA-phaseI-phaseII%2Fdb%2FHMP-16S-I-II%3F&gbGridXAttr=seq_center&gbGridYAttr=primer_region_PLUS_body_site&xlabel=seq_center&ylabel=primer_region_PLUS_body_site&gridTitle=Samples%20from%20HMP-16S-I-II&pageTitle=Sample%20Grid%20Viewer:%20Samples%20from%20HMP-16S-I-II

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Microbiome Tutorial1. Introduction to 16S Analysis

• Genboree.org/microbiome– http://genboree.org/theCommons/projects/pub-gen-microbiome/wiki/Microbiome_Tutorial

» 5 Stool samples» 5 Throat samples

2. Replicating Published Data• A core gut microbiome in obese and lean twins (Turnbaugh et al, Nature, 2009)• Study objective: To address the question of how host genotype,

environmental exposures, and host adiposity influence the gut microbiome – http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2677729/pdf/nihms74182.pdf

» 49 Lean Stool samples» 45 Obese Stool samples

– Comparing the alpha diversity between lean and obese stool samples

3. Utilizing HMP control data– > 13,000 samples– 18 body sites– 3 primer regions

4. KEGG Mashups– Subset of HMP WGS HUMAnN output

» 50 samples» 7 body sites

Aleksandar Milosavljevic, Baylor College of Medicine amilosav@bcm.edu

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KEGG Mashup

• Genes and Pathways– View samples + tracks within Grid Viewer– View output within Gene Browser and Pathway

Browser– View Pathways within KEGG

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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KEGG Mashup

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Questions?

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Virtual Integration

• Accessing data that exists within different physical servers

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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Virtual Integration

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• http://genboree.org/java-bin/sampleGridViewer.jsp?dbList=http%3A%2F%2Fgenboree.org%2FREST%2Fv1%2Fgrp%2FHMP-16S-rRNA-phaseI-phaseII%2Fdb%2FHMP-16S-I-II%3F,http%3A%2F%2Fgenboree.org%2FREST%2Fv1%2Fgrp%2FPublic_16S_experiment_data%2Fdb%2FDisease_X%3F&gbGridXAttr=TYPE&gbGridYAttr=DNA_extraction_site_PLUS_seq_center_PLUS_body_site_PLUS_primer_region&xlabel=TYPE&ylabel=DNA_extraction_site_PLUS_seq_center_PLUS_body_site_PLUS_primer_region&gridTitle=Samples%20from%20HMP-16S-I-II,Disease_X&pageTitle=Sample%20Grid%20Viewer:%20Samples%20from%20HMP-16S-I-II,Disease_X

• http://genboree.org/java-bin/sampleGridViewer.jsp?dbList=http%3A%2F%2Fgenboree.org%2FREST%2Fv1%2Fgrp%2FHMP-16S-rRNA-phaseI-phaseII%2Fdb%2FHMP-16S-I-II%3F,http%3A%2F%2Fgenboree.org%2FREST%2Fv1%2Fgrp%2FPublic_16S_experiment_data%2Fdb%2FDisease_X%3F&gbGridXAttr=primer_region&gbGridYAttr=body_site&xlabel=primer_region&ylabel=body_site&gridTitle=Samples%20from%20HMP-16S-I-II,Disease_X&pageTitle=Sample%20Grid%20Viewer:%20Samples%20from%20HMP-16S-I-II,Disease_X

• http://genboree.org/java-bin/sampleGridViewer.jsp?dbList=http%3A%2F%2Fgenboree.org%2FREST%2Fv1%2Fgrp%2FHMP-16S-rRNA-phaseI-phaseII%2Fdb%2FHMP-16S-I-II%3F,http%3A%2F%2Fgenboree.org%2FREST%2Fv1%2Fgrp%2FPublic_16S_experiment_data%2Fdb%2FDisease_X%3F&gbGridXAttr=barcode&gbGridYAttr=primer_region_PLUS_body_site&xlabel=barcode&ylabel=primer_region_PLUS_body_site&gridTitle=Samples%20from%20HMP-16S-I-II,Disease_X&pageTitle=Sample%20Grid%20Viewer:%20Samples%20from%20HMP-16S-I-II,Disease_X

• http://genboree.org/java-bin/sampleGridViewer.jsp?dbList=http%3A%2F%2Fgenboree.org%2FREST%2Fv1%2Fgrp%2FHMP-16S-rRNA-phaseI-phaseII%2Fdb%2FHMP-16S-I-II%3F,http%3A%2F%2Fgenboree.org%2FREST%2Fv1%2Fgrp%2FPublic_16S_experiment_data%2Fdb%2FDisease_X%3F&gbGridXAttr=TYPE&gbGridYAttr=primer_region_PLUS_body_site&xlabel=TYPE&ylabel=primer_region_PLUS_body_site&gridTitle=Samples%20from%20HMP-16S-I-II,Disease_X&pageTitle=Sample%20Grid%20Viewer:%20Samples%20from%20HMP-16S-I-II,Disease_X