Northern blot

Northern blot is a technique used in molecular biology for detection of specific RNA (Ribonucleic acid) sequences in a sample. The technique was developed by James Alwine, David Kemp, and George Stark at Stanford University in 1977. It is named after the Southern blot, a method for DNA detection developed by Edwin Southern. While the Southern blot is used for DNA, the Northern blot is used for RNA.

Principle

The Northern blot method involves the separation of RNA samples by gel electrophoresis, transfer of the RNA from the gel to a membrane (blotting), and detection of specific RNA sequences on this membrane. The RNA sequences are detected by hybridization to a labeled complementary probe.

Procedure

The procedure of Northern blotting includes several steps:

1. Isolation of RNA: The first step in Northern blotting is the isolation of total RNA from the sample. This can be done using various methods, including phenol extraction or the use of commercial kits.
2. Gel electrophoresis: The RNA samples are then separated by size using gel electrophoresis. The gel used is usually an agarose gel, and the RNA is denatured before loading to ensure that it runs according to its size.
3. Transfer to membrane: The separated RNA is then transferred from the gel to a membrane. This is done by capillary action, where the liquid containing the RNA is drawn up through the gel and onto the membrane.
4. Hybridization: The membrane is then incubated with a labeled probe that is complementary to the RNA sequence of interest. The probe will bind to its complementary sequence on the membrane, allowing for the detection of this specific RNA sequence.
5. Detection: The final step is the detection of the probe, which indicates the presence of the RNA sequence of interest. This can be done using various methods, depending on the type of label used on the probe.

Applications

Northern blotting is used in various fields of molecular biology, including:

• Studying gene expression: Northern blotting can be used to study the expression of specific genes, by detecting the RNA produced from these genes.
• Virus detection: Northern blotting can be used to detect specific viral RNA sequences, allowing for the identification of viral infections.
• Cancer research: Northern blotting can be used in cancer research to study changes in gene expression that occur in cancer cells.

See also

• Southern blot
• Western blot
• Eastern blot

Northern blot Resources
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