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Chromatin Immunoprecipitation and the Chip on Chip technique.

Chromatin Immunoprecipitation and the Chip on Chip technique. Fredrik Fagerström Billai B E A- Core Facility for B ioinformatics and E xpression A nalysis. Department of Biosciences & Nutrition Karolinska Institutet.

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Chromatin Immunoprecipitation and the Chip on Chip technique.

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  1. Chromatin Immunoprecipitation and theChip on Chip technique. Fredrik Fagerström Billai B E A- Core Facility for Bioinformatics and Expression Analysis. Department of Biosciences & Nutrition Karolinska Institutet

  2. Agenda-General introduction to Chromatin and Transcriptional and Epigenetic control.-Chromatin Imunnoprecipitation, tiling arrays and the ”Chip on Chip” technique Fredrik Fagerström Billai , BEA

  3. Chromatin Fredrik Fagerström Billai , BEA

  4. Images of Chromatin A = 30nm fiber of an interphase chromosome B = Nucleosomes along a strand of DNA Fredrik Fagerström Billai , BEA

  5. The Chromatin Subunit model Fredrik Fagerström Billai , BEA

  6. The Nucleosome Crystal Structure Fredrik Fagerström Billai , BEA

  7. Histones are modified at Specific Amino Acids Histone Code hypothesis - Modifications of the Histone tails act as marks that can be read by other proteins to control the expression or replication of chromosomal regions. The coding in the histones may be heritable. Fredrik Fagerström Billai , BEA

  8. Specific modifications are associated with specific functions. Fredrik Fagerström Billai , BEA

  9. Methylation of DNA Transcriptional gene silencing Chromatin compaction Genome stability X chromosome inactivation (females) Fredrik Fagerström Billai , BEA

  10. Fredrik Fagerström Billai , BEA

  11. Eukaryotic mRNAs are synthesized by the Pol II complex Fredrik Fagerström Billai , BEA

  12. Gene regulation in the eukaryotes Wasserman and Sandelin (2004) Fredrik Fagerström Billai , BEA

  13. Chromatin Immunoprecipitation Fredrik Fagerström Billai , BEA

  14. Chip- General outline Fredrik Fagerström Billai , BEA

  15. Chip analysis of ERα recruitment to estrogen responsive gene promoters. Ray et al. 2006 Fredrik Fagerström Billai , BEA

  16. Chip Quantification Calella et al. 2007 Fredrik Fagerström Billai , BEA

  17. Real Time PCR quantification Fredrik Fagerström Billai , BEA

  18. Real Time PCR quantification Fredrik Fagerström Billai , BEA

  19. Chip on Chip Outline Fredrik Fagerström Billai , BEA

  20. Genome Wide Chip Solutions Classic Two Color Arrays Affymetrix Tiling Arrays Illumina Deep Sequencing Fredrik Fagerström Billai , BEA

  21. * * * * * Affymetrix array architecture Single stranded, labeled ‘target’ GeneChipProbe Array Oligonucleotide ‘probe’ Millions of copies of a specific oligonucleotide probe 1.28cm 1.28cm Fredrik Fagerström Billai , BEA

  22. Affymetrix genechip family Fredrik Fagerström Billai , BEA

  23. Tiling Arrays Fredrik Fagerström Billai , BEA

  24. Tiling arrays provide high resolution for identifying bound fragments Overlapping 25-mer fragments Boyer et al. 2005 Fredrik Fagerström Billai , BEA

  25. Quantile Normalization Fredrik Fagerström Billai , BEA

  26. Identifying bound genomic fragments by Intensity Percentile Ranking. Fredrik Fagerström Billai , BEA

  27. Integrated Genome Browser, IGB Fredrik Fagerström Billai , BEA

  28. Genome-wide profiling of PRC1Polycomb chromatin binding in Drosophila melanogaster. Fredrik Fagerström Billai , BEA

  29. Chip profiles Fredrik Fagerström Billai , BEA

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