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. 2016 Nov 21;215(4):575–590. doi: 10.1083/jcb.201607043

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© 2016 Smoyer et al.

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Figure 1. — Split-GFP to study protein localization. (A) Schematic of the GFP complementation system. Protein molecular masses in kilodaltons based on amino acid composition. (B) Schematics show the GFP₁₁-mCherry-Pus1 reporter and INM protein Asi1-GFP_1–10, which were expressed alone (top and middle) or together in yeast. Fluorescence in the red (561-nm) and green (488-nm) channels is shown. Residual background is autofluorescence, which was shown by multispectral imaging (not depicted). (C) Schematic illustrating the localization of GFP₁₁-mCherry reporters to detect signal inside the nucleus (GFP₁₁-mCherry-Pus1), ONM/ER surface (GFP₁₁-mCherry-Scs2TM), ER lumen (mCherry-Scs2TM-GFP₁₁), and cytoplasm (GFP₁₁-mCherry-Hxk1). An ONM/ER protein containing the Scs2 transmembrane domain fused to GFP_1–10 on its cytoplasmic side is also shown. (D) Example images of reporters alone (top) or with GFP_1–10-Scs2TM (bottom). Bars, 2 µm.