FIGURE 3.

(A) Processing reaction of pre-miR-138-2 into mature miR-138 by recombinant Dicer (rDcr) resolved on a 15% denaturing PAGE. (B) As in A. The addition of increasing amounts of HeLa cytoplasmic extract effectively abolishes the processing of pre-miR-138-2. The band corresponding to the respective pre-miRNA is indicated by “pre,” whereas the band representing the mature miRNA is designated by “m.” (C) Processing of pre-miR-19a into mature miR-19a is not abolished by addition of increasing amounts of HeLa cytoplasmic extract. The efficiency of processing is reduced by the addition of HeLa cytoplasmic extracts; however, the level of processing does not decrease with higher amounts of HeLa cytoplasmic extracts. (D) HeLa cells were transfected with the indicated amounts of a plasmid encoding the sequence of pre-miR-124a. The conversion of pre-miR-124a into mature miR-124a was monitored by Northern blotting. (E) HeLa cells were transfected with the indicated amounts of a plasmid encoding the sequence of pre-miR-138-2. In contrast to pre-miR-124a, HeLa cells failed to process pre-miR-138-2 into mature miR-138. Note that pre-miR-138-2 is more abundant than pre-miR-124a.