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. 2017 Oct 19;7:13605. doi: 10.1038/s41598-017-13997-7

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© The Author(s) 2017

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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Genetic elements of the doxycycline-regulated Nkkc1 suppression system. The construct for Actin-tTS::Nkcc1 ^tetO/tetO. Nkcc1 consists of 26 exons. The translation initiation site (ATG) is situated at exon 1, and the stop site marks the end of translation at exon 26. The tetracycline operator site (tetO) was inserted just upstream of ATG to form Nkcc1 ^tetO/tetO. Nkcc1 ^tetO/tetO homozygotes behave like wild-type mice. The tetracycline trans-silencer (tTS) tethers tetO in the absence of doxycycline (DOX), resulting in the suppression of Nkcc1 expression. In the presence of DOX (green circles), tTS-mediated gene knockdown does not occur. Gray boxes: coding regions in exons 1 and 26.