FIG 3.

Possible restoration of the growth phenotype by complementing the NcGRX5 gene in ΔNcGRX5 parasites. (A) Construction strategy for complementation parasites (comΔNcGRX5) and overexpression parasites (NcGRX5 OE). (B) Confirmation of NcGRX5 OE and comΔNcGRX5 parasites by PCR. (C) Confirmation of NcGRX5 OE and comΔNcGRX5 parasites by Western blotting using mouse anti-HA antibody. Actin was used as a loading control. The immunoblotting band was quantitatively evaluated by ImageJ based in two independent experiments. The error bars represent the standard error. (D) Confirmation of NcGRX5 OE and comΔNcGRX5 parasites by IFA. The location of NcGRX5 (green) was visualized with anti-HA, and SRS2 (red) was used as a parasite shape marker. (E) Comparing the overall growth ability of WT, ΔNcGRX5, NcGRX5 OE, and iΔNcGRX5 parasites using plaque assays. (F) Measurements of plaque areas using the pixel point in Photoshop C6S software (Adobe, USA). The data are derived from three independent experiments. (G) Comparison of the invasion ability of comΔNcGRX5, ΔNcGRX5, NcGRX5 OE, and wild-type (WT) parasites. (H) Comparison of the intracellular replication ability of comΔNcGRX5, ΔNcGRX5, NcGRX5 OE, and WT parasites. (I) Survival rate of mice after infection with 1 × 10⁷ comΔNcGRX5, ΔNcGRX5, NcGRX5 OE, and WT parasites. DHFR, dihydrofolate reductase.