Figure 7. Emi2 Is Expressed Post-meiotically.

(A and B) Real-time PCR analysis was performed to analyze mRNA levels of genes at different ages

(A) and in elutriation fractions (B) of testes. Expression values were normalized to house-keeping gene Elongation Factor 2 (EF2) and expressed relative to P7 (A) or Fractions 1 + 2 (B). Sequences of primers used are listed in Table S2. Mean values obtained from testes extracted from three mice per age group are reported in (A). For (B), results are representative of two independent elutriation experiments, with testes from three mice used per experiment. Error bars represent mean ± SD.

(C) Protein extracts of testes collected from control or Emi2KO mice at various ages were subjected to SDS-PAGE and western blotting with the indicated antibodies. Results are representative of three independent experiments.

(D) Protein extracts of fractions collected from testicular cell elutriation were subjected to SDS-PAGE and western blotting with the indicated antibodies. Fraction 1: residual bodies + elongated spermatids; Fraction 2: residual bodies + elongated spermatids; Fraction 3: round spermatids; Fraction 4: round spermatids + spermatocytes; and Fraction 5: spermatocytes. Results are representative of three independent experiments.

(E) Testes were isolated from control or Emi2KO mice (n = 3) at P20 and processed for immunohistochemistry with antibodies against Emi2. Scale bars: 100 μm.

(F) Testes from control or Emi2KO mice (n = 3) at P60 were processed for immunofluorescent staining with antibodies against Emi2.

(G) Cauda epididymal sperm from wild-type mice (n = 3) were immunostained for Emi2 and SP56.

(H) Human testicular tissues were immunostained for Emi2.

White arrowheads point to spermatids with acrosomal staining. Results are representative of three independent experiments.