Microarray Data Analysis

Microarray Data Analysis Microarray Data Analysis Stuart M Brown NYU School of Medicine proteinRNADNA transcription translation replication The Central Dogma of Molecular Biology DNA is transcribed in[.]

Microarray Data Analysis Stuart M Brown NYU School of Medicine The Central Dogma of Molecular Biology DNA is transcribed into RNA which is then translated into protein DNA transcription RNA translation protein replication Measured by Microarray What is a Microarray • A simple concept: Dot Blot + Northern • Reverse the hybridization - put the probes on the f ilter and label the bulk RNA • Make probes for lots of genes - a massively parallel experiment • Make it tiny so you don’t need so much RNA from your experimental cells • Make quantitative measurements Microarrays are Popular  At NYU Med Center we are now collecting about GB of microarray data per week (60 chips, 6-10 different experiments)  PubMed search "microarray"= 13,948 papers 2005 2004 2003 2002 2001 2000 = = = = = = 4406 3509 2421 1557 834 294 5000 4500 4406 4000 3509 3500 3000 2500 2421 2000 1557 1500 1000 500 834 294 2000 2001 2002 2003 2004 2005 A Filter Array DNA Chip Microarrays • Put a large number (~100K) of cDNA sequences or synthetic DNA oligomers onto a glass slide (or other subtrate) in known locations on a grid • Label an RNA sample and hybridize • Measure amounts of RNA bound to each square in the grid • Make comparisons – Cancerous vs normal tissue – Treated vs untreated – Time course • Many applications in both basic and clinical research cDNA Microarray Technologies • Spot cloned cDNAs onto a glass microscope slide – usually PCR amplif ied segments of plasmids • Label RNA samples with dif ferent colors of f lourescent dye - control vs experimental • Mix two labeled RNAs and hybridize to the chip • Make two scans - one for each color • Combine the images to calculate ratios of amounts of each RNA that bind to each spot Spot your own Chip (plans available for free from Pat Brown’s website) Robot spotter Ordinary glass microscope slide Combine scans for Red & Green False color image is made from digitized fluorescence data, not by superimposing scanned images