*4.1. Plant Materials*

The cassava plants South China 124 (SC124) were kindly provided by Dr. Wei Hu (Institute of Tropical Bioscience and Biotechnology, Haikou, China). Two-week-old tissue culture cassava plants (SC124) were transferred to small pots and grown in a chamber for two weeks until the experiment at 25 °C under a 16 h light/8 h dark cycle.

## *4.2. Vectors and Vector Construction*

The pEGAD and pBI121 vectors were used to express *GFP* and *GUS*, respectively. The full-length sequences of *GFP* and *GUS* were driven by the 35S promoter. The VIGS assay used pTRV1 and pTRV2 vectors, which have been described previously [52]. The partial sequence of the *MePDS* gene was cloned into the multiple cloning site of the pTRV2 vector, and the primers are listed in Supplementary Table S1.

## *4.3. Agrobacterium Infiltration of Cassava*

*Agrobacterium* strains GV3101 and AGL-1 were transformed by different plasmids: pEGAD, pBI121, pTRV1, pTRV2, and pTRV2-*MePDS*, respectively. GV3101 was cultivated in liquid LB medium containing 50 mg/L kanamycin, 20 mg/L rifampicin, and 50 mg/L gentamycin, while AGL-1 was cultivated in LB medium with 50 mg/L kanamycin, 20 mg/L rifampicin, and 50 mg/L carbenicillin. Both were then shook at 28 °C at 200 rpm for 2 days. The bacteria was centrifuged at 4000 rpm for 10 min, then the supernatant was discarded. The remaining bacteria was washed with double-distilled water, then centrifuged, and the supernatant was discarded again. The bacterial sediment was resuspended in MMA solution (10 mM MgCl, 10 mM MES, and 150 µM acetosyringone), the OD<sup>600</sup> was adjusted to 1, and then the resuspended bacterial solution was placed in the dark for 3 h. For the transient expression assay, the standing bacterial solution was infiltrated into the second and third leaves from the top of the cassava by a 1 mL needle. For the VIGS assay, the bacterial solutions containing pTRV1 or pTRV2-*MePDS* were mixed with the same volume, and the mixture of pTRV1 or pTRV2 was used as the mock. The mixed solution was infiltrated into both the leaves and axillary buds of cassava plants to keep the silencing effect [48]. After infiltration, the cassava plants were removed to the chamber at the same light and temperature.

## *4.4. DNA and RNA Extraction*

DNA was extracted by the Plant Genomic DNA Extraction Kit (DP305, TIANGEN, Beijing, China). Total RNA was extracted by the RNAprep Pure Plant Kit (Polysaccharides & Polyphenolics-rich) (DP441, TIANGEN, Beijing, China). The remaining DNA from the extracted RNA was digested by

RNase-free DNase I (EN0521, Thermo, Waltham, MA, USA). Then, the quality and concentration of DNA and RNA were examined by Nano Drop 2000 (Thermo, Waltham, MA, USA).
