MBIO 4612 Lecture Notes - Lecture 17: Tata-Binding Protein, Hela, Chromatin
9 Feb 2020
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Document Summary
Allows analysis of in vivo interactions of chromatin-associated proteins with specific regions of genomic dna. Can use ab against specifically acetylated histones. Un-crosslink dna, purify & assay via pcr. Experiment viral infection induces pattern of histone acetylation at ifn-b promoter. Ifn-b promoter was detected via pcr with promoter-specific primers. Cross-linked chromatin was immunoprecipitated with specific ab. Looking for transcription of interferon by looking at patterns of acetylation on each histone. Non-histone rows: input dna = dna present in all columns, ifn-b mrna = shows transcription was occurring, a-tbp = shows tbp recruitment (tata box binding protein) No band = lower levels of acetylation. Note similarity between h3 k14 acetylation, tbp recruitment, & transcription. Chip assay + microarrays examines differences between tested & control dna. Purified dna from chip is mixed with ab against modified histone or chromatin proteins: basically, want to bind the dna with a protein that can be detected (fluorescent probe, etc. )
