Northern blotting is a laboratory technique used in molecular biology to study gene expression
It is used to detect and quantify RNA molecules in a sample, providing information about gene expression levels.
Involves gel electrophoresis to separate RNA molecules by size, followed by transfer to a membrane and detection using specific probes.
RNA samples are denatured (usually with formaldehyde) and loaded onto an agarose gel. Electrophoresis separates RNA molecules based on size.
After electrophoresis, RNA molecules are transferred ("blotted") from the gel to a membrane, usually nylon or nitrocellulose.
RNA on the membrane is then immobilized by UV cross-linking or baking, making it accessible for probe hybridization.
Specific RNA molecules of interest are detected using labeled DNA or RNA probes that hybridize with complementary sequences on the membrane.
Once hybridized, the probe is detected using methods such as autoradiography (for radioactive probes) or chemiluminescence (for non-radioactive probes).
The intensity of the signal corresponds to the amount of RNA present in the sample, allowing for quantification of gene expression levels.
Used to analyze RNA size, detect alternative splicing, assess gene expression changes under different conditions, and validate results from other gene expression profiling techniques.
Provides information about specific RNA molecules, relatively straightforward technique once established, and allows for both qualitative and quantitative analysis of gene expression.