Figure 2.

Analyses of miRNAs and ta-siRNAs in ΔPpDCL4 Mutants.

(A) Summed, scaled, repeat-normalized sRNA abundances for all MIRNA-derived sRNAs from wild-type and ΔPpDCL4. Units are normalized reads per 10 000.

(B) Expression analysis of miR390, miR166, miR156, and miR160 in protonema tissue by RNA gel blots. U6 snRNA served as control for normalization. Numbers under blots indicate normalized expression values with respect to wild-type.

(C) Chart indicating the expression of TAS3a-d derived sRNAs inferred from sequencing data of wild-type and ΔPpDCL4.

(D) RNA gel blots for ta-siRNAs derived from TAS3a and TAS3c precursors in wild-type and ΔPpDCL4 mutants. U6snRNA served as control for normalization. Numbers under blots indicate normalized expression values for off-sized ta-siRNAs with respect to the 21-nt wild-type ta-siRNAs.

(E) Transcript levels of ta-siRNA target genes (PpARF; Phypa_203442 and PpAP2; Phypa_65352) in wild-type (WT) and ΔPpDCL4 mutants determined by qRT–PCR. Expression levels were normalized to the constitutively expressed control gene PpEF1α.

(F) RLM 5' RACE products of the miRNA and ta-siRNA target PpARF from WT and ΔPpDCL4 mutants. Arrows mark products of the expected size (1: miRNA-mediated cleavage product; 2: ta-siRNA-mediated cleavage product) that were eluted, cloned, and sequenced. Numbers above miRNA:target and ta-siRNA:target alignments indicate sequenced RACE products with the corresponding 5' end.