Figure 5.

Depurination of tobacco ribosomes in vivo. Ribosomes were isolated from wild-type tobacco plants and 50 μg of ribosomes were treated with 300 ng of PAP as described. RNA was extracted (TR) and divided in half, and one half was treated with aniline (TR + A). Total RNA (20 μg) isolated from transgenic plants expressing PAP-v (26139), the active site mutant (144-7), the C-terminal deletion mutant (145-13), and wild-type tobacco plants (WT) was divided in half, and one half was treated with aniline (+ A). RNAs were separated on a 4.5% urea/polyacrylamide gel and visualized after staining with ethidium bromide (Upper) or were separated on a 1.3% formaldehyde agarose gel and probed with a PCR fragment complementary to the 3′ end of the rRNA (Lower). The position of the 360-nt diagnostic cleavage product of the rRNA is indicated with an arrow.