2.4 Differentially expressed proteins (DEP) analysis
TMT-based quantitative proteomic analysis was the same as description in our previous article, including Protein extraction and digestion, protein digestion, TMT labeling, low-pH nano-HPLC-MS/MS analysis, and the database searching (Zhang et al. 2019). Database searching was conducted using the present RNA-seq results (Accession codes: SUB7765255) by Mascot.
For protein quantitation, a protein was required to contain at least two unique peptides. 9 protein ratios were produced by comparing 3 M4 and 3 M0 (i.e., M4a/M0a, M4a/M0b, M4a/M0c, M4b/M0a, M4b/M0b, M4b/M0c, M4c/M0a, M4c/M0b, M4c/M0c). then the P values were determined using ANOVA (Student’s t-test) and the 9 ratios. The mean comparisons of pairs among the 9 protein ratios calculated fold changes. Finally proteins withP < 0.05 and fold change > 1.2 were considered to be DEP (Chen et al.2013b; Zhang et al. 2019).