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. 2020 Sep 2;10:14437. doi: 10.1038/s41598-020-71474-0

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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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Distribution of reconstituted GFP is dependent on GRAPHIC probe molecular structure. A red nucleus (labeled by H2B-mCherry) indicates an NT cell, and a blue nucleus (labeled by H2B-Azurite) indicates a CT cell. (A) GRAPHIC: NT probe containing an acidic leucine zipper domain (LZA) and CT probe containing a basic leucine zipper domain (LZB). Reconstituted GFP was observed only at the boundary between NT and CT cells. White box indicates position of higher magnification view. (B) dGRAPHIC: Leucine zipper domain was removed from both NT and CT probes. dGRAPHIC showed almost all reconstituted GFP signals on NT cells, compared with CT cells. (C) A probe pair consisting of NT and CT probes with 3 spacer domains (15 aa × 3) inserted between the GFP-CT fragment and GPI anchor domain. Reconstituted GFP was equally observed on both NT and CT cells. (D) Combination of NT and CT probes with 4 spacer domains (15 aa × 4). Reconstituted GFP was detected predominantly on CT cells. Scale bar, 40 µm and for higher magnification 20 μm.