〔Abstract〕 Objective To study the effect of EFHD2 protein deletion in Sertoli cells on Occludin, a component of tight junction protein and the localization and expression of EF-hand domain family member D2(EFHD2) in mouse testis. Methods Total RNA and protein were extracted from adult miceʼs heart, liver, spleen, lung, kidney, brain and testis tissues. The mRNA and protein levels of EFHD2 in each organ tissue were detected by qRT-PCR and Western blot. Immunofluorescence and immunohistochemistry detected the localization and expression of EFHD2 in testicular tissues. SiRNA interference was used to reduceEFHD2in Sertoli cells to detect Occludin protein expression. Results qRT-PCR showed that the expression ofEFHD2was the highest in the testis. Western blot results showed that the expression level increased in testis tissue. Indirect immunofluorescence and immunohistochemistry results showed that the protein was mainly distributed in Sertoli cells and co-localized with cytoskeletal Vimentin, indicating that the protein was expressed in Sertoli cells.After the decrease of EFHD2 protein expression, Occludin protein expression also decreased. Conclusion The expression ofEFHD2protein in the testis is relatively high, mainly distributed in Sertoli cells of the testis, co-localized with Vimentin, and can affect the normal expression of tight junction protein Occludin. It is suggested that EFHD2 can promote and maintain the junction structure of Sertoli cells and provide a stable microenvironment for spermatogenesis.