GeneExpression-M.TevfikDORAKsWebsite.ppt

Complex assemblies of proteins control eukaryotic transcription A variety of regulatory proteins interact with DNA and each other Gene Expression in Prokaryotes Three kinds of RNA MIAME Guidelines “Minimum Information About a Microarray Experiment” /Workgroups/MIAME/miame.html Provides a minimum standard that should be followed to objectively interpret findings from array experiments and ensure reproducibility of results Guidelines are provided for: Experimental design Samples used and preparation Hybridization techniques Microarray protocol Processing and Analysis of Data Davidson University (Microarray Animation) /courses/genomics/chip/chip.html Imagecyte (Microarray Animation) /array2.html Microarray Data Analysis (Microarray Bibliography) /microarray Nigel Walker, NIEHS (www) Traditional gene expression analysis:Northern Blotting Northern blotting detects specific RNAs RNA is isolated from cells and separated using electrophoresis probed with radioactive cDNA from a specific gene Method can be used to determine steady-state level of a transcript in a specific RNA mixture Serial analysis of gene expression (SAGE) ? 9 to 11 base “tags” correspond to genes ? measure of gene expression in different biological samples ? SAGE tags can be compared electronically Serial analysis of gene expression (SAGE) Page 169 SAGENet: /findings/index.html DNA FOOTPRINTING ANALYSIS RIBONUCLEASE PROTECTION ASSAY NUCLEASE PROTECTION ASSAY In this example, an mRNA containing a point mutation (indicated by the inverted triangle in the mRNA on the right) is distinguished from its normal, non-mutated counterpart (mRNA on the left). The mRNA is mixed with a single-stranded 32P-labeled DNA or RNA probe that (1) has sequences perfectly complementary to the nonmutated region of interest in the mRNA, and (2) extends for some length beyond the mRNA. The mixture is heated then cooled to allow the probe to anneal to its complementary sequences in the mRNA. The annealed mixture is t