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. 2020 Nov 10;10:19396. doi: 10.1038/s41598-020-76538-9

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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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/.

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Effects of Dux family overexpression on the change in gene expression pattern in the early 2- and 4-cell stages. cRNA encoding Dux, DuxG1, DuxG9, or firefly luciferase (Luc, control) were microinjected into a blastomere of a 2-cell stage embryo at 16–20 hpi and cultured in vitro. Cells were collected and RT-PCR was conducted to amplify Dux family, Zfp352, Zscan4d, Galk1, Wsb1 and rabbit α-globin (external control) at 40 hpi. MII stage oocytes and non injected embryos at 16 and 40 hpi were also collected for RT-PCR. Thirty cells were used for each sample. (A) Electrophoresis images of PCR products. (B–F) The band densities in (A) were quantified using image J. The band density of each gene is relative to that of rabbit α-globin. The values at the early 2-cell stage (16 hpi) were set to 1 and the relative values were calculated. Five (Dux family, Zfp352 and Zscan4d) and three (Galk1 and Wsb1) independent experiments were conducted. Error bars represent SE. Asterisks indicate significant difference (p < 0.05) by paired Student t-test.