Gene expression in scRNA-seq and quantitative real-time PCR. (A) Differential gene expression in scRNA-seq. Single trophozoites of axenic culture, CHO incubation, and in vivo groups are marked in orange, green, and blue, respectively. (B) qRT-PCR assays of CS, MGL, Hgl, Igl-1, Igl-2, CP2, CP5, and AP-A genes of trophozoites cultured under normal or cysteine-deficient conditions. c, medium contained normal concentration of cysteine; w/o, medium without cysteine. The gene expression levels are represented using the 2−ΔΔCT of the target gene relative to the β-actin gene. The y axes are linear coordinates, and numbers correspond to the fold increase over 1.0 given to the axenic culture group. *, P < 0.05; **, P < 0.01; ***, P < 0.001.