Fig. 3. Identification of kinetochore-targeting domain of 53BP1 by GFP-fusion analysis. (A) The various portion of m53BP1 that were fused to GFP and tested for their ability to be recruited (+) or not (–) to kinetochores (K.L.), and to localise (+) or not (–) to the nucleus during interphase (N.L.). (B) Prometaphase HeLa cells expressing GFP fused to the indicated m53BP1 amino acid position were stained with monoclonal antibody against CENP-E to visualise kinetochores. CENP-E was detected with Texas Red-conjugated anti-mouse secondary antibodies. Chromosomes were stained with DAPI. GFP:m53BP1^1-1957 (a), GFP:m53BP1^753-1957 (b), GFP:m53BP1^1-1601 (d), GFP:m53BP1^753-1601 (f), GFP:m53BP1^1140-1703 (g), GFP:m53BP1^1140-1601 (h), GFP:m53BP1^1169-1601 (i), GFP:m53BP1^1220-1601 (j) properly generated a GFP signal colocalising with CENP-E, whereas GFP:m53BP1^1294-1957 (c), GFP:m53BP1^1-1139 (e), GFP:m53BP1^1294-1601 (k), GFP:m53BP1^1220-1515 (l) failed to do so. Images were recorded with a conventional microscope.