Am Af

+ Protein probe

+ Calmodulin probe (d)

+ Calmodulin probe (d)

+ Protein probe

+ Protein probe

▲ EXPERIMENTAL FIGURE 15-4 Protein microarray analysis can reveal protein-protein and protein-lipid interactions. High-level expression cloning was used to produce 5800 yeast proteins, which were purified and then spotted in duplicate on microscope slides. Some 13,000 protein samples can be spotted in half the area of a standard microscope slide. A probe was prepared by covalently attaching biotin to calmodulin or to phosphatidylinositol triphosphate (IP3). The yeast proteome chips were incubated with the biotinylated probe, and then excess probe was washed off. Calmodulin or IP3 bound to proteins in the microarray was detected with fluorescence-labeled streptavidin, a bacterial protein that strongly and specifically binds biotin. (a) A part of a proteome chip probed to reveal the location and approximate amounts of all proteins, which are spotted in duplicate columns. (b) The same preparation probed with biotinylated calmodulin. The two green signals correspond to a calmodulin-binding protein spotted in duplicate. (c) A different field of protein spots. (d) IP3 binds to two of the proteins in (c), each in duplicate. [Courtesy of Paul Bertone, Yale University.]

criterion, however, is the phenotype of cells or animals lacking the gene. Knowledge of the functions of genes and relations among them during development can be obtained by systematically eliminating the function of each gene that is normally expressed in a tissue, one at a time, and observing what goes wrong. This procedure is now theoretically possible in C. elegans, which is particularly susceptible to RNA interference (RNAi). In this technique, delivery of double-stranded RNA into worms by injection, feeding, or soaking leads to the destruction of the corresponding endogenous mRNA (see Figure 9-43). The phenotype of worms that develop from RNAi-treated embryos is often similar to the one that would result from inactivation of the corresponding gene itself. In initial studies with C. elegans, RNA interference with 16,700 genes (about 86 percent of the genome) yielded 1722 visibly abnormal phenotypes. The genes whose

Your Heart and Nutrition

Your Heart and Nutrition

Prevention is better than a cure. Learn how to cherish your heart by taking the necessary means to keep it pumping healthily and steadily through your life.

Get My Free Ebook


Post a comment