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. 2018 Oct 18;8:15433. doi: 10.1038/s41598-018-33032-7

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© The Author(s) 2018

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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Targeting vector construction strategies. (a) The top graph shows the wild-type allele with exon 1 to 5 in the murine iGb3S WT locus, together with relevant enzyme restriction sites and probes for Southern blot analysis. The middle graph shows the targeting vector constructed by replacing a 1.5-kb fragment encoding the iGb3S exon 5 with a loxP-flanked neomycin-resistance gene cassette (neo), and a diphtheria toxin A (DTA). The lower graph shows the mutated allele with an11.4 kb Nde1-fragment at the 5′-arm immediately upstream of the neo cassette and 12.3 kb of the Nde1-fragment at the 3′-arm immediately downstream of the neo cassette. Coding and non-coding regions of exon 1–5 are marked by filled and non-filled boxes, respectively. Homology arms are depicted as bold lines. (b,c) show Southern blotting results. (b) Genomic DNA was digested with NdeI in the 5′-probe and resulted in 25.2 kb and 11.4 kb fragments. (c) Genomic DNA was digested with NdeI in the 3′-probe and resulted in 25.2 kb and 12.3 kb fragments. M, DNA marker; lanes 1 and 2 are mut/mut, lanes 3 and 4 are mut/wt; lanes 5 and 6 are wt/wt.