Figure 3. The impact of mutations on CFAP52 expression.
(A) Western blotting analysis showed that CFAP52 expression was not detected in HEK293T cells transfected with the mutant-CFAP52c.1128G>A plasmid. (B) Electrophoresis showed a decrease in the molecular weight of the PCR products generated from mutant CFAP52c.203G>T (263 bp) compared with WT CFAP52 (400 bp) in the minigene experiment. (C) Sanger sequencing of cDNA of the splicing mutation showed the deletion of exon 2 in the c.203G>T variant clone. (D) The pattern diagram depicts the adverse effects caused by the CFAP52 splicing mutation c.203G>T. (E) Western blotting analysis showed that HEK293T cells transfected with the mutant CFAP52c.203G>T plasmid did not express CFAP52. (F) Immunofluorescence staining showed that CFAP52 expression in the patient’s spermatozoa was absent compared with that of a healthy control (blue, DAPI; green, CFAP52; red, α-tubulin). Scale bars, 5 μm. All experiments were repeated three times with similar results.