FIGURE 3.

(A) Conditional KO of TbAGO1 results in up-regulation of large transcripts homologous to the Ingi retroposon transcripts. Total RNA was prepared from cells grown in the presence or absence of tet for the times indicated above each lane. Ingi transcripts were detected with an Ingi probe as described (Janzen et al. 2006). Control: hybridization to the calflagin mRNAs. (B) Conditional KO of TbAGO1 results in an inhibition of Ingi siRNA accumulation. TbAGO1KO_c cells were incubated in the presence (+) or absence (−) of tet for 24 h. Total RNA was prepared and enriched for small RNAs as described previously (Shi et al. 2004b). The small RNAs were separated on a denaturing 15% polyacrylamide gel, electroblotted onto a nylon membrane, and hybridized with a sense radiolabeled riboprobe representing approximately the first 1000 nt of the Ingi coding region. Control shows a group of cross-hybridizing bands. The position of a 26-nt DNA marker is shown by a bar on the side.