Nevertheless, differential processing of stem-loop precursors, small RNA duplexes, and, in general, miRNA biochemical properties challenge expression analysis methods usually used to analyze mRNAs.
Moreover, the population of short sequence reads produced allows the identification of differential processing of stem-loop precursors by DICER-LIKE1, and mature miRNA isoforms within and between species.
To do this, we used a set of "detector" models, testing selectively for the processing of stems, inflected forms and phrases (Fig. 3).
In addition, promoter transcripts also lack motifs important for the processing of the stem, such as TG dinucleotide at the base of the stem and CNNC motif downstream of the stem, as described in Auyeung et al.
Preferential accumulation of the loop mRNA, however, indicates that degradation of the hairpin transcript occurs mainly via processing of the stem of the hairpin.
In wild type plants, the major pathway for degradation of the ΔGUS-SUG hairpin transcript appears to be DCL4-dependent processing of the stem into primary siRNAs.
DCL4-dependent processing of the stem of the hairpin into primary siRNAs is the major pathway utilized in wild type and dcl2 mutant plants.
When both DCL2 and DCL4 are defective, degradation again occurs via processing of the stem into primary siRNAs, but this time involves DCL3 and possibly DCL1.
Suppression of hairpin and sense transgene silencing by the P1/HC-Pro and P38 viral suppressors was associated with elimination of secondary siRNA accumulation, but the suppressors did not block processing of the stem of the hairpin transcript into primary siRNAs.
When both DCL2 and DCL4 are inactive, silencing is again dependent on primary siRNAs, and processing of the stem of the hairpin by DCL3 becomes evident (Figures 2B and 3B).
