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Printable Handouts
Navigable Slide Index
- Intro slide
- The target gene problem (1)
- Example 1: the E2F family of transcription factors
- Example 2: the zinc finger transcription factors
- The target gene problem (2)
- Do all family members regulate the same genes?
- How can we find target genes?
- Standard assays to identify target genes
- The ChIP assay
- Controls for ChIP assays
- Identification of in-vivo binding sites
- Cloning/sequence analysis of ChIP fragments
- PCR amplification of candidate promoters
- How can one identify candidate target promoters?
- Combining gene expression profiling and ChIP
- Targets may be missed due to primer location
- Analyzing ChIP samples using microarrays
- ChIP-on-chip analyses (1)
- Amplicon verification
- ChIP-on-chip analyses (2)
- Types of Microarray Platforms
- ChIP-on-chip analyses (3)
- Analysis of candidate promoters
- High throughput analysis of candidate promoters
- Analysis of genomic tiling array (1)
- Combining microarray analyses
- What is a CpG microarray?
- How do you obtain CpG islands?
- Creation of a CpG island library
- Experimental design for ChIP-CpG
- How do you analyze CpG arrays?
- Identification of positive CpG islands
- Experimental design
- Analysis of genomic tiling arrays (2)
- ChIP-on-chip analyses: promoter arrays
- Core promoter arrays
- Tiled core promoter arrays
- ChIP-on-chip analyses: whole genome arrays
- Genomic tiling arrays: screening an entire genome
- Example of mapping E2F1 binding sites
- Experimentally determined/consensus sites
- Some consensus E2F sites are bound by E2F1
- The location of the E2F site determines occupancy
- Confirmation steps
- Confirmation using amplicons
- Confirmation using a biological replicate
- Have you identified a bona fide target gene?
- Does factor binding regulate the target gene?
- E2F6 siRNA
- E2F6 knockdown affects gene expression
- Summary
- Advantages of ChIP-on-chip
- Limitations of the current technology
- Websites for detailed protocols
Topics Covered
- Identifying genomic targets of DNA-binding factors
- Experimental methods
- Chromatin immunoprecipitation (ChIP) assay
- Probing microarrays
- ChIP-on-chip assays for comprehensive analysis
- Data interpretation and follow-up studies
Talk Citation
Farnham, P. (2007, October 1). Genome-wide analyses of protein-DNA interactions [Video file]. In The Biomedical & Life Sciences Collection, Henry Stewart Talks. Retrieved December 12, 2024, from https://doi.org/10.69645/LRMO5475.Export Citation (RIS)
Publication History
Financial Disclosures
- Prof. Peggy Farnham has not informed HSTalks of any commercial/financial relationship that it is appropriate to disclose.