BIOL3150 Lecture Notes - Polyacrylamide Gel Electrophoresis, Electrophoresis, Proteinase K
Document Summary
Functional genomics part ii: protein-dna interaction assays and the encode (chapter 11: study protein-dna interactions by, emsa (electrophoretic mobility shift) assay or gel shift assay. Used to identify which dna segment binds to a protein or protein complex. Used to identify region of dna occupied by a protein complex. How: end labeled restriction fragments combined with nuclear extract and without, proteins bind to dna with nuclear extract, limited. Dnase i digestion (cannot cut where protein), so will have gap where proteins bound (footprint), will know protein binding somewhere in this region. Can couple with ngs to identify dnase i hypersensitivity for entire genome (peak = greater accessibility to dna region), footprints correspond to transcription factor binding sites. Can be combined with microarray technology to map essentially all: chromatin immunoprecipitation (chip) genomic binding sites of a dna-binding protein (chip-chip) Rna polymerase recognizes -35 box and unravels dna near -10 box.
