The Topic The Problem The Idea The Results Computational Analysis of Methylome Sequencing Data Master Thesis Bioinformatics Till Helge Helwig Eberhard-Karls-University Tübingen Wilhelm-Schickard-Institut für Informatik & Max Planck Institute for Developmental Biology February 22, 2011 Computational Analysis of Methylome Sequencing Data Till Helge Helwig 1 / 21
Outline The Topic 1 What is a Methylome? Why is the Methylome Interesting? The Problem 2 Obtaining the Methylome via Sequencing Problems with the Common Approach The Idea 3 How Can Computer Science Help? Evaluated Methods The Results 4 Performance Comparison What Do the Results Imply?
Outline The Topic 1 What is a Methylome? Why is the Methylome Interesting? The Problem 2 Obtaining the Methylome via Sequencing Problems with the Common Approach The Idea 3 How Can Computer Science Help? Evaluated Methods The Results 4 Performance Comparison What Do the Results Imply?
The Topic The Problem The Idea The Results What is a Methylome? The Methylome Entirety of methylated nucleotides (e.g. cytosines) in the DNA Addition of a methyl group converts cytosine into 5-methylcytosine NH 2 NH 2 N N N O N O H H Computational Analysis of Methylome Sequencing Data Till Helge Helwig 4 / 21
The Topic The Problem The Idea The Results What is a Methylome? The Methylome Entirety of methylated nucleotides (e.g. cytosines) in the DNA Addition of a methyl group converts cytosine into 5-methylcytosine NH 2 NH 2 N N N O N O H H Computational Analysis of Methylome Sequencing Data Till Helge Helwig 4 / 21
m m m A CG T A CG T A CG T MT T GC A T GC A T GC A m m A CG A CG T T MT m MT m T GC T GC A A m m A CGT A CGT A CGT m MT T GC A T GC A T GC A m m m The Topic The Problem The Idea The Results What is a Methylome? Properties of the Methylome Additional layer of information within the DNA Computational Analysis of Methylome Sequencing Data Till Helge Helwig 5 / 21
m m m A CG T A CG T A CG T MT T GC A T GC A T GC A m m A CG A CG T T MT m MT m T GC T GC A A m m A CGT A CGT A CGT m MT T GC A T GC A T GC A m m m The Topic The Problem The Idea The Results What is a Methylome? Properties of the Methylome Additional layer of information within the DNA Methylations are created by methyltransferases Computational Analysis of Methylome Sequencing Data Till Helge Helwig 5 / 21
The Topic The Problem The Idea The Results What is a Methylome? Properties of the Methylome Additional layer of information within the DNA Methylations are created by methyltransferases Maintenance of methylations after transcription m m m A CG T A CG T A CG T MT T GC A T GC A T GC A m m A CG A CG T T MT m MT m T GC T GC A A m m A CGT A CGT A CGT m MT T GC A T GC A T GC A m m m Computational Analysis of Methylome Sequencing Data Till Helge Helwig 5 / 21
The Topic The Problem The Idea The Results What is a Methylome? Properties of the Methylome Additional layer of information within the DNA Methylations are created by methyltransferases Maintenance of methylations after transcription m m m A CG T A CG T A CG T MT T GC A T GC A T GC A m m A CG A CG T T MT m MT m T GC T GC A A m m A CGT A CGT A CGT m MT T GC A T GC A T GC A m m m Environmental factors influence the methylome Computational Analysis of Methylome Sequencing Data Till Helge Helwig 5 / 21
The Topic The Problem The Idea The Results What is a Methylome? Properties of the Methylome Additional layer of information within the DNA Methylations are created by methyltransferases Maintenance of methylations after transcription m m m A CG T A CG T A CG T MT T GC A T GC A T GC A m m A CG A CG T T MT m MT m T GC T GC A A m m A CGT A CGT A CGT m MT T GC A T GC A T GC A m m m Environmental factors influence the methylome The methylome is highly variable ... ...between different species Computational Analysis of Methylome Sequencing Data Till Helge Helwig 5 / 21
The Topic The Problem The Idea The Results What is a Methylome? Properties of the Methylome Additional layer of information within the DNA Methylations are created by methyltransferases Maintenance of methylations after transcription m m m A CG T A CG T A CG T MT T GC A T GC A T GC A m m A CG A CG T T MT m MT m T GC T GC A A m m A CGT A CGT A CGT m MT T GC A T GC A T GC A m m m Environmental factors influence the methylome The methylome is highly variable ... ...between different species ...between organisms of the same species Computational Analysis of Methylome Sequencing Data Till Helge Helwig 5 / 21
The Topic The Problem The Idea The Results What is a Methylome? Properties of the Methylome Additional layer of information within the DNA Methylations are created by methyltransferases Maintenance of methylations after transcription m m m A CG T A CG T A CG T MT T GC A T GC A T GC A m m A CG A CG T T MT m MT m T GC T GC A A m m A CGT A CGT A CGT m MT T GC A T GC A T GC A m m m Environmental factors influence the methylome The methylome is highly variable ... ...between different species ...between organisms of the same species ...between different cell types of the same organism Computational Analysis of Methylome Sequencing Data Till Helge Helwig 5 / 21
The Topic The Problem The Idea The Results Why is the Methylome Interesting? Transcription Inhibition Methylated nucleotides can inhibit the transcription Computational Analysis of Methylome Sequencing Data Till Helge Helwig 6 / 21
The Topic The Problem The Idea The Results Why is the Methylome Interesting? Transcription Inhibition Methylated nucleotides can inhibit the transcription Relevance for different research fields: Developmental biology (e.g. for association studies) Medicine (e.g. for tumorgenesis) Ecology (e.g. documentation of environmental changes) ... Computational Analysis of Methylome Sequencing Data Till Helge Helwig 6 / 21
Outline The Topic 1 What is a Methylome? Why is the Methylome Interesting? The Problem 2 Obtaining the Methylome via Sequencing Problems with the Common Approach The Idea 3 How Can Computer Science Help? Evaluated Methods The Results 4 Performance Comparison What Do the Results Imply?
U A GT A C T G A U A m C A GT A C T G A C A bisulfite G T C A T GAC T G T treatment G T C A T GA U T G T m A T C A T GA C T A T forward complement R C T A GT A C TG A T A P forward G T C A T GA T T G T reverse complement C A GT A CT A A C A reverse complement synthesis methylated unmethylated 0:2:2:0 1:1:1:1 100% CG 50% CG The Topic The Problem The Idea The Results Obtaining the Methylome via Sequencing Making the Methylome Visible Standard sequencing can not identify methylations Computational Analysis of Methylome Sequencing Data Till Helge Helwig 8 / 21
The Topic The Problem The Idea The Results Obtaining the Methylome via Sequencing Making the Methylome Visible Standard sequencing can not identify methylations Bisulfite treatment makes methylations visible: U A GT A C T G A U A m C A GT A C T G A C A bisulfite G T C A T GAC T G T treatment G T C A T GA U T G T m A T C A T GA C T A T forward complement R C T A GT A C TG A T A P forward G T C A T GA T T G T reverse complement C A GT A CT A A C A reverse complement synthesis methylated unmethylated 0:2:2:0 1:1:1:1 100% CG 50% CG Computational Analysis of Methylome Sequencing Data Till Helge Helwig 8 / 21
The Topic The Problem The Idea The Results Obtaining the Methylome via Sequencing Making the Methylome Visible Standard sequencing can not identify methylations Bisulfite treatment makes methylations visible: U A GT A C T G A U A m C A GT A C T G A C A bisulfite G T C A T GAC T G T treatment G T C A T GA U T G T m A T C A T GA C T A T forward complement R C T A GT A C TG A T A P forward G T C A T GA T T G T reverse complement C A GT A CT A A C A reverse complement synthesis methylated unmethylated 0:2:2:0 1:1:1:1 100% CG 50% CG Sequencing now reports only methylated cytosines Computational Analysis of Methylome Sequencing Data Till Helge Helwig 8 / 21
The Topic The Problem The Idea The Results Obtaining the Methylome via Sequencing Sequencing Protocol Bisulfite treatment inserted into the sequencing protocol tissue sample bisulfite treatment sequencing reads DNA extraction PCR mapping DNA double strand Solexa sequencing reads per position Computational Analysis of Methylome Sequencing Data Till Helge Helwig 9 / 21
The Topic The Problem The Idea The Results Obtaining the Methylome via Sequencing Sequencing Protocol Bisulfite treatment inserted into the sequencing protocol tissue sample bisulfite treatment sequencing reads DNA extraction PCR mapping DNA double strand Solexa sequencing reads per position Methylation rates calculated from the read counts per position Computational Analysis of Methylome Sequencing Data Till Helge Helwig 9 / 21
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