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The northern blot is a technique used in molecular biology research to study gene expression. It takes its name from its similarity to the Southern blot technique, named for biologist Edwin Southern and used to study DNA. The major difference is that RNA, rather than DNA, is analyzed in the northern blot. Both techniques use electrophoresis and detection with a hybridization probe. The northern blot technique was developed in 1977 by James Alwine, David Kemp, and George Stark at Stanford University.[1]
The gels may be run on either agarose or denaturing polyacrylamide, the latter being preferable for smaller fragments of RNA. Unlike in the Southern blot, formaldehyde is added to the gel and acts as a denaturant to agarose. For polyacrylamide, urea is the denaturant.
As in the Southern blot, the hybridization probe may be made from DNA or RNA.
A variant of the procedure known as the reverse northern blot is occasionally used. In this procedure, the substrate nucleic acid (that is affixed to the membrane) is a collection of isolated DNA fragments, and the probe is RNA extracted from a tissue and radioactively labelled.
The use of DNA microarrays that have come into widespread use in the late 1990s and early 2000s is more akin to the reverse procedure, in that they involve the use of isolated DNA fragments affixed to a substrate, and hybridization with a probe made from cellular RNA. Thus the reverse procedure, though originally uncommon, enabled northern analysis to evolve into gene expression profiling, in which many (possibly all) of the genes in an organism may have their expression monitored.
References
- ^ Alwine JC, Kemp DJ, Stark GR (1977). "Method for detection of specific RNAs in agarose gels by transfer to diazobenzyloxymethyl-paper and hybridization with DNA probes". Proc. Natl. Acad. Sci. U.S.A. 74 (12): 5350–4. doi:. PMID 414220.
See also
External links
- Northern Blot analysis of mRNA from mammalian polyribosomes (a protocol)
- Northern blot protocols with variations from many labs
- Northern Blot A great northern blot resource complete with diagram and protocol.
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