Sara is an undergraduate student who is doing an internship in the research laboratory described above. Just before Sara started working in the lab, the restriction map above was made of the 47-kb NotI restriction fragment containing the prion-protein gene (distances between restriction sites are in kb).
Since smaller DNA fragments cloned into plasmids are more easily analyzed than large DNA fragments cloned into BACs, Sara has been asked to “subclone” the 6.1-, 10.5-, 4.1-, and 8.2-kb BamHI DNA fragments containing the prion-protein gene into the pUC19 plasmid vector. Her mentor gives her some intact pUC19 plasmid DNA, some of the purified 47-kb NotI fragment, and shows her where the lab’s stocks of DNA ligase and BamHI are stored. Describe the steps Sara should take to complete her task. In your answer, address how she will identify plasmids that contain genomic DNA inserts, and how she will verify that she has identified clones containing each of the desired genomic BamHI fragments.
20. Imagine that you have been able to clone the structural gene for an enzyme in a catecholamine biosynthesis pathway from the adrenal gland of rats. How could you use this cloned DNA as a probe to determine whether this same gene functions in the rat brain?
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