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. 2021 Jun 1;12:3184. doi: 10.1038/s41467-021-23378-4

Fig. 2. Spermatogenesis was impaired in Zfp541 knockout male.

Fig. 2

a The allele with targeted deletion of Exon1–13 in Zfp541 gene was generated by the introduction of CAS9, the synthetic gRNAs designed to target Exon1 and the downstream of Exon13 (arrowheads), and ssODN into C57BL/6 fertilized eggs. Arrows: PCR primers for genotyping. Four lines of KO mice were established. Line #16 of Zfp541 KO mice was used in most of the experiments, unless otherwise stated. b Immunoblot analysis of testis extracts prepared from mice with the indicated genotypes (P21). Biologically independent mice (N = 2) were examined in two separate experiments. c Testes from Zfp541± and Zfp541 KO (8-weeks old). Testis/body-weight ratio (mg/g) of Zfp541± and Zfp541 KO mice (8-weeks old) is shown below (mean with SD). n: the number of animals examined. Statistical significance is shown by **p = 0.0021 (two-tailed t-test). Scale bar: 5 mm. d Hematoxylin and eosin staining of the sections from WT, Zfp541±, and Zfp541 KO testes (8-weeks old). Biologically independent mice (N = 3) for each genotype were examined. Scale bar: 50 μm. e Hematoxylin and eosin staining of the sections from WT, Zfp541±, and Zfp541 KO epididymis (8-weeks old). Biologically independent mice (N = 3) for each genotype were examined. Scale bar: 50 μm. f Seminiferous tubule sections (8-weeks old) were stained for SYCP3, ZFP541, and DAPI. pL: preleptotene, Pa: pachytene spermatocyte, rS: round spermatid, eS: elongated spermatid. Boundaries of the seminiferous tubules are indicated by white dashed lines. Roman numbers indicate the seminiferous tubule stages. Biologically independent mice (N = 3) for each genotype were examined. Scale bars: 15 μm.