Table S1. The primers used in this study. from A fungal milRNA mediates epigenetic repression of a virulence gene in <i>Verticillium dahliae</i> JinYun ZhaoJian-Hua ZhaoPan ZhangTao WangSheng GuoHui-Shan 2018 MiRNAs in animals and plants play crucial roles in diverse developmental processes under both normal and stress conditions. miRNA-like small RNAs (milRNAs) identified in some fungi remain functionally uncharacterized. Here, we identified a number of milRNAs in <i>Verticillium dahliae</i>, a soil-borne fungal pathogen responsible for devastating wilt diseases in many crops. Accumulation of a <i>V. dahliae</i> milRNA1, named VdmilR1, was detected by RNA gel blotting. We show that the precursor gene <i>VdMILR1</i> gene is transcribed by RNA polymerase II and is able to produce the mature VdmilR1, in a process independent of <i>V. dahliae</i> DCL (Dicer-like) and AGO (Argonaute) proteins. We found that an RNaseIII domain-containing protein, VdR3, is essential for <i>V. dahliae</i> and participates in VdmilR1 biogenesis. VdmilR1 targets a hypothetical protein-coding gene, <i>VdHy1</i>, at the 3′UTR for transcriptional repression through increased histone H3K9 methylation of <i>VdHy1</i>. Pathogenicity analysis reveals that <i>VdHy1</i> is essential for fungal virulence. Together with the time difference in the expression patterns of VdmilR1 and <i>VdHy1</i> during fungal infection in cotton plants, our findings identify a novel milRNA, VdmilR1, in <i>V. dahliae</i> synthesized by a noncanonical pathway that plays a regulatory role in pathogenicity and uncover an epigenetic mechanism for VdmilR1 in regulating a virulence target gene.This article is part of the theme issue ‘Biotic signalling sheds light on smart pest management’.