Northern blot analysis

Southern blot analysis except with RNA instead of DNA, and RNA is short enough that it doesn't need to be digested

RNA sep by electrophoresis (sep by size), transferred to memb, probed

negatives: length process, low sensitivity/large sample req, only 1 gene per hybridization

Polymerase Chain Reaction

PCR

DNA (NOT RNA--can use reverse transcriptase on mRNA), primers (dT), mixture of nucleotides and DNA polymerase, buffers, thermocycling

1. Denaturation 2. Annealing 3. Extension 4. Repeat to amplify DNA

high sensitivity, only small sample needed

negative: not quantitative

Real time PCR

quantitative

use conventional PCR setup, add Syber Green dye, moniter reaction-->graph shows change over time

threshold of detection.  Farther right-->less starting material

in situ hybridization

localization of DNA or mRNA in cell to study spatial characteristics

not quantitative

shows where a gene is being expressed

starting material=tissue

DNA microarray analysis

method for simulataneous eval of multiple genes

comparisons between two populations of cells (normal/diseased, control/treated)

Yellow/red(overexpression)/green(underexpression)

negatives: $$$, data overload, only good for known genes/products

Differential expression analysis by differential display

compare two different RNA populations

differential display--PCR uses random primers, shows diff expression of mult genes btw 2 samples

positive: large # of differentially expressed genes can be shown on one gel

negative: large # of primers needed