Quantifying Arabidopsis Guvermectin Response

Two-week-old Arabidopsis thaliana leaves were sprayed with 50 mg L⁻¹ guvermectin (GV) once, and the whole plant was ground with liquid nitrogen at 0, 1, 3, 5, and 7 d after GV treatment, then total RNA was extracted using TRIzol reagent [74 (link)]. Meanwhile, the plants sprayed with 0 mg L⁻¹ GV were used as a control at each point. For qRT-PCR, 1 μg total RNA per sample was treated with the PrimeScript™ RT reagent Kit with gDNA Eraser (RR047A, Takara, San Jose, CA, USA). qRT-PCR was performed with SYBR Master Mix (Q711-03, Vazyme, Nanjing, China) on the Bio-Rad iQ5 optical system software (Bio-Rad, Hercules, CA, USA). UBQ5 was used as the internal control gene for expression level normalization. The transcript level of each gene was calculated using the double ΔCt method [34 (link)]. Data analysis was conducted in GraphPad Prism 8. The primers were summarized in Table S3.