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. 2019 Jun 27;9:9339. doi: 10.1038/s41598-019-45411-9

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© The Author(s) 2019

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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Effects of ZNF518B expression on the level of EMT markers. DLD1 cells were treated with mixed ZNF518B siRNAs or with scrambled siRNA (control) for 48 and 72 h and the EMT markers were analysed. The levels of the SNAI1 (a) and CHD1 (b) genes were determined by RT-qPCR. (c) Western blots showing the level of ZNF518B (left) and N-cadherin and SNAIL (right) after 72 h of silencing ZNF518B. Histone deacetylase 1 (HDAC) or glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were used as loading controls. Due to the different size of ZNF518B and of the EMT markers, the electrophoreses were carried out separately in 6% and 12% polyacrylamide gels, respectively. Ponceau-stained membranes were cropped according to the molecular weight of the corresponding proteins and the resulting strips were developed with the antibodies described under Materials and Methods. (d) ImageJ analysis of the western blots of panel c. Four grey values relative to the loading controls were measured in every case and averaged. Statistical analysis was carried out with the Mann-Whitney test. (*p < 0.05; ***p < 0.001).