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| Modern Genetic Analysis Chapter 10. Recombinant DNA Technology Using Cloned DNA Figure 10-18. Gel electrophoresis and blotting in identifying specific cloned genes. RNA or DNA restriction fragments are applied to an agarose gel and electrophoresed. The various fragments migrate at differing rates according to their respective sizes. The gel is placed in buffer and covered by a nitrocellulose filter and a stack of paper towels. The fragments are denatured to single strands, so that they can stick to the filter. They are carried to the filter by the buffer, which is wicked up by the towels. The filter is then removed and incubated with a radioactively labeled single-stranded probe that is complementary to the targeted sequence. Unbound probe is washed away, and X-ray film is exposed to the filter. Because the radioactive probe has hybridized only with its complementary restriction fragments, the film will be exposed only in bands corresponding to those fragments. Comparison of these bands with labeled markers reveals the number and size of the fragments in which the targeted sequences are found. This information can be used to position the sequences along restriction maps. This procedure is termed Southern blotting when DNA is transferred to nitrocellulose and Northern blotting when RNA is transferred. (From J. D. Watson, M. Gilman, J. Witkowski, and M. Zoller, Recombinant DNA, 2d ed. Copyright © 1992 by Scientific American Books.)

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