## 2 Results
### Expression of CYP81A and miRNAs Targeting Their Transcripts  
Results concerning cytochromes P450 belonging to the 81A subfamily and miRNAs targeting their transcripts are reported in Figure 1. Significant changes in the expression levels (fold change > 2) of genes and miRNAs are identified with a \(p\) < 0.05. As shown in Figure 1A, after profoxydim spraying, in the SUS biotype the expression of _CYP81A12_ decreased by half (from 1.00 \(\pm\) 0.05 to 0.45 \(\pm\) 0.03), while that of _osa-miRR64-5p_ increased (from 1.06 \(\pm\) 0.46 to 1.79 \(\pm\) 0.22). Meanwhile, gene expression significantly in  
Figure 1: Expression levels of the miRNAs studied (red) and the mRNA of their target genes (blue) in susceptible (SUS) and resistant (RES1 and RES2) biotypes of _Echinochlo oryzae1o_ before (BT) and after (AT) profoxydim treatment. Mean and standard error values are indicated as bar plots and error bars, respectively. The expressions of genes and miRNAs were normalized with those of the respective internal references (_b-Actin_ and _non-coding small nuclear RNA U6_). (**A**) _CYP81A12_ and _osa-miRR64-5p_; (**B**) _CYP81A21_ and _osa-miRR64-5p_; (**C**) _CYP81A22_ and _ath-miRR396b-5p_; (**D**) _CYP81A24_ and _osa-miRR2099-5p_; (**E**) _CYP81A6_ and _osa-miRR395f_; (**F**) _CYP81A63_ and _osa-miRR396a-5p_. _t-_test: \(\textbf{\emph{r}}\) \(p\) < 0.05.  
creased in RES1 (from 1.86 \(\pm\) 0.21 to 8.58 \(\pm\) 1.31) and RES2 (from 1.14 \(\pm\) 0.03 to 2.32 \(\pm\) 0.09), while _osa-miR166a-5p_ transcription decreased. As shown in Figure 1B, after herbicide application, _CYP8 IA21_ expression remained almost equal in the SUS biotype, while it significantly increased in RES1 (0.45 \(\pm\) 0.05 vs. 2.75 \(\pm\) 0.48) and in RES2 (0.006 \(\pm\) 0.001 vs. 2.00 \(\pm\) 0.01). The expression of _osa-miR166d-5p_, which displayed similar values before treatment (\(\sim\)1.00) in all biotypes, remained almost equal in SUS, while significantly it decreased after profoxydim administration in RES1 and RES2. As shown in Figure 1C, after herbicide spraying, the expression of _CYP8 IA22_ mRNA slightly rose in the SUS biotype (1.01 \(\pm\) 0.16 vs. 1.24 \(\pm\) 0.20), while it significantly increased in RES1 (16.35 \(\pm\) 2.17 vs. 100.10 \(\pm\) 13.42) and in RES2 (7.97 \(\pm\) 1.08 vs. 40.87 \(\pm\) 3.69). _Ath-miR396b_ transcription decreased in all biotypes after treatment. A similar result was observed concerning _CYP8 IA24_ and _osa-miR2099-5p_ expression (Figure 1D). After herbicide application, gene expression slightly increased in the SUS biotype (1.03 \(\pm\) 0.33 vs. 1.39 \(\pm\) 0.44), while it significantly increased in both RES1 (20.04 \(\pm\) 7.69 vs. 129.18 \(\pm\) 53.58) and RES2 (24.12 \(\pm\) 2.31 vs. 100.02 \(\pm\) 11.19). _osa-miR2099-5p_ transcription was therefore found not to be triggered by profoxydim spraying in SUS, RES1, and RES2. The expression of _CYP8 IA6_ (Figure 1E) increased in SUS (1.25 \(\pm\) 0.82 vs. 1.46 \(\pm\) 0.97) and significantly rose in the RES1 (from 3.87 \(\pm\) 2.22 to 25.80 \(\pm\) 13.67) and RES2 (form 0.85 \(\pm\) 0.29 to 2.09 \(\pm\) 0.79) biotypes after herbicide spraying. Meanwhile, _osa-miR8396a-5p_ transcription displayed almost equal values in the SUS biotype before and after profoxydim administration, while it decreased in the RES1 and RES2 biotypes. _CYP8 IA63_ expression and _osa-miR8396a-5p_ transcription after herbicide treatment (Figure 1F) displayed profiles similar to those analyzed above: gene expression rose in SUS (1.00 \(\pm\) 0.09 vs. 1.37 \(\pm\) 0.79) and significantly increased in RES1 (0.63 \(\pm\) 0.12 vs. 3.94 \(\pm\) 0.64) and in RES2 (0.33 \(\pm\) 0.05 vs. 3.17 \(\pm\) 0.74), while _osa-miR8396a-5p_ transcription decreased.