Comparative Proteomic Analysis of Plant Samples

Total latex proteins were extracted as described (Wang et al., 2010 (link)). Protein concentration was determined by Bradford assay with BSA as a standard. For 2-DE, 1,300 μg proteins were loaded onto 24-cm, pH 3-10 linear gradient IPG strips (GE Healthcare, Uppsala, Sweden), and isoelectric focusing and gel electrophoresis were performed as described (Wang et al., 2013 (link)). Three biological replicates were performed for each sample. Protein spots in gel were visualized by the CBB-G250 staining, and images were analyzed with the ImageMaster 2D Platinum software (GE Healthcare). The average Vol% for each matched protein spot was calculated based on all the detected gel images from the three biological repeats (Supplementary Figure S1). First, we compared Eth, Mo, and EMo with CK, as well as EMo with Eth and EMo with Mo; the protein spots that, in at least one pair, showed an at least 1.5-fold change in abundance (p < 0.05) were termed as differentially expressed protein (DEP) spots. Then, these protein spots were positively identified by MALDI TOF MS/MS.