Synchronized worms were collected from the plates, washed at least three times with M9 buffer, and resuspended in TransZol Up reagent (TransGen Biotech, #ET111-01). Worm samples were frozen in liquid nitrogen and freeze-thawed for at least five times. Total RNA was isolated by chloroform extraction followed by isopropanol precipitation, washed with 75% (v/v) ethanol and dissolved in RNase-free water. Small RNA libraries for small RNA seq were prepared by BGI Tech using DNBSEQ small RNA seq protocols.