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. 2017 Dec 19;7:17771. doi: 10.1038/s41598-017-17651-0

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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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Drug selection-free RMCE. LPV-encoded GFP reporter genes in CHO cells were replaced by the DV_RMCE sequences. (a) Representative fluorescence microscopy images and flow cytometry plots of, from left to right, the CHO.S18 master clone, the unselected RMCE pool after the RMCE transfection, an isolated RFP⁺ clone (CHO.S18-R) and the same clone after Cre-mediated RFP gene deletion (CHO.S18-RΔRFP). Scale bar = 100 µm. (b) Verification of the expected RMCE (left) and deletion events (right) by PCR analysis of genomic DNA. The nomenclature of the cell lines and pools is according to Fig. 2a, above. Shown on the left is a multiplex PCR with 3 primers where the lower band of 537 bp is indicative of the intact LPV, whereas the upper band of 1,329 bp is indicative of the RMCE reaction. Shown on the right is a PCR analysis of integrated DV_RMCE, before (upper band of 4,571 bp) and after (lower band of 2,157 bp) Cre/loxP deletion. For details of the PCR patterns refer to suppl. Fig. S9. (M: Hyperladder 1 kb). The uncropped gel pictures are shown in S11a.