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Figure 4 | Reproductive Biology and Endocrinology

Figure 4

From: Spam1-associated transmission ratio distortion in mice: Elucidating the mechanism

Figure 4

Spam1 transcripts which are compartmentalized are absent from the bridges and are associated with the cytoskeleton. a) EM autoradiography of seminiferous tubules after in situ hybridization with a tritiated (3H-labeled) Spam-1 antisense RNA probe. Note that silver grains are associated with the ER (arrowheads) but not with any other major spermatid structures such as the chromatoid bodies (A), the radial bodies (B) or the microtubules of the manchette (C). D is an intercellular bridge where the curvatures at the top and bottom represent the outer limits of the bridge. While some grains are seen in association with the ER in the vicinity, they are absent from the bridge. The circles centered over the silver grains include profiles of ER (arrowheads). (E) Late spermatids (S) and Sertoli cells (Se) are unreactive. (F) Shows the cytoplasm of round spermatid (RS) step 8 of a control section hybridized to a sense probe. Co, chromatoid body; Rb, radial body; M, manchette. X19,000 b. Northern hybridization of Spam1 and β-actin mRNAs in free cytosolic-, cytoskeletal-, and membrane-bound testicular RNA fractions. The fractions were separated by subcellular fractionation techniques. A) shows Northern blotting, while B) shows total RNA as a loading control with ethidium bromide staining. The presence of cytoskeletal-bound β-actin RNA in the free cytosolic fraction suggests that Spam1 in the latter could be present as a contaminant due to the preparation procedure.

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