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. 2020 Mar 24;10:5335. doi: 10.1038/s41598-020-62091-y

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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No fertilization was obtained with Spaca6 KO sperm in vitro. Fertilization rate (FR) (mean ± SEM) following cumulus-intact IVF assay at 10⁶ sperm per ml or zona-free IVF assay at 10⁵ sperm per ml for 3 hours. No fertilization was obtained with Spaca6 KO sperm in C57BL/6 (a) and FVB/N (b) lines. While no differences concerning the FR were observed when comparing the WT and heterozygous groups in C57BL/6 background (a,c), significant ones were obtained in FVB/N background both in cumulus-intact (b) and zona-free assays (d) (P < 0.0001). Regarding the fertilization index (FI; mean ± SEM), measurable only in zona-free IVF assay, and that gives the average number of fused sperm per oocyte observed after insemination, significant differences were observed in both backgrounds (e for C57BL/6 and f for FVB/N) when heterozygous groups were compared to WT ones (P < 0.0001). The sperm and oocyte Spaca6 genotypes are indicated below each figure.