基因组信息学9.pdf

ChIP-Seq What is ChIP-Seq?  Chromatin-Immunoprecipitation (ChIP)- Sequencing  ChIP - A technique of precipitating a protein antigen out of solution using an antibody that specifically binds to the protein.  Sequencing – A technique to determine the order of nucleotide bases in a molecule of DNA.  Used in combination to study the interactions between protein and DNA. Workflow of ChIP-sequencing  Part 1: ChIP  ChIP is a powerful method to selectively enrich for DNA sequences bound by a particular protein in living cells.  The ChIP process enriches specific crosslinked DNA- protein complexes using an antibody against a protein of interest.  The antibodies may be attached to a solid surface, may have a magnetic bead ChIP-seq work flow ChIP-seq video ChIP-on-chip ChIP-seq VS ChIP-on-chip ChIP-Seq Analysis Pipeline ChIP-Seq: Genome Alignment  Several Aligners Available  BWA  NovoAlign  Bowtie  Currently the Sequencing analysis pipeline uses the Stampy as the default aligner for all sequencing.  All aligner output containing information about the mapping location and quality of the reads are out put in SAM format Single End  Flow cell clusters (reads) represent strand-specific fragment ends  A sequenced fragment with reference strand sequence  ACCTAACCGAAC can give rise to only 2 possible 4bp reads:  ACCT and GTTC (assuming no sequencing errors)  A double-stranded DNA fragment can be represented in set of reads by 0, 1, or 2 of its ends  Paired End data : a fragment is represented by 0 or 2 of its ends  A sequenced fragment with reference strand sequence  ACCTAACCGAAC will give rise to exactly 2 4bp reads:  ACCT and GTTC (assuming no sequencing errors) Types of Events Base Calling Basics  Discover interaction sites from aligned reads  Idea: loci with lots of reads/fragments = signal site  But loc