Northern blot

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The northern blot is a technique used in molecular biology research to study gene expression. It takes its name from the similarity of the procedure to the Southern blot procedure, named for biologist Edwin Southern, used to study DNA, with the key difference that, in the northern blot, RNA, rather than DNA, is the substance being analyzed by electrophoresis and detection with a hybridization probe. This 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 gels depending on the size of the RNA to be detected. A notable difference in the procedure in case of agarose gels, (as compared with the Southern blot) is the addition of formaldehyde which acts as a denaturant. For smaller fragments denaturing polyacrylamide urea gels are employed.

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 was occasionally (although, infrequently) 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 the one-at-a-time study of gene expression using northern analysis to evolve into gene expression profiling, in which many (possibly all) of the genes in an organism may have their expression monitored.

[edit] 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:10.1073/pnas.74.12.5350. PMID 414220.