Figure 2.

siRNA detection by RNase Protection Assay (RPA). Synthetic mC9 siRNA (c9si) and different sources of RNA were probed by RPA (top) using a mouse caspase 9 anti-sense probe. The positions of intact ³²P-labeled probe (~200 nt), siRNAs, xylene cyanol (XC, ~60 bp) and bromophenol blue (BPB, ~15 bp) are indicated. tR (yeast tRNA); Gsi (GAPDH synthetic siRNA); c9si (single mC9 synthetic siRNA); Tc9 1 and 2 (total RNA samples from two Tc9 plants); WT (total RNA from wt plant); Dc9 (Dicer-derived mC9 siRNA); BL (body-labeled Dicer siRNA). Bands marked Δ are artifacts of the RPA as described in the text. Quantitation of Dc9si (bottom) was achieved by comparison with known amounts of synthetic siRNAs following resolution on 15% non-denaturing PAGE gels and staining with EtBr.