Fig 1: Minimal co-localization of MATR3 with lamin A/C or histones. Mouse C2C12 cells were grown on glass cover slips and transiently transfected with pEF.Bos vectors encoding MATR3:YFP variants noted on the figure. At 24 hours post-transfection, the cells were fixed, immunostained with antibodies to lamin A/C (Sigma-Aldrich #SAB4200236) or histone H3 (Abcam #ab8898) and imaged. The images shown are representative of images seen in 3 independent transfections, viewing 30 to 50 cells on each cover slip. Each image is from a single z-plane captured on a confocal microscope (see Methods). Nikon elements software was used to determine the extent of interaction between MATR3 and lamin A/C or histone H3 (Supplemental Figures S16 and S17). MATR3 shows limited co-localization with either lamin A/C or histone H3.
Fig 2: Mitotic chromosome binding is correlated with TF-DNA co-localization in interphase. a Immunofluorescence labeling of H3K9me3 (Abcam, #ab8898) and Hoechst staining of a NIH-3T3 nucleus. Scale bar: 3 µm. b Automatic detection of regions displaying different densities of DNA: Heterochromatic (circled in red), DNA-rich (between red and yellow circles), or DNA-poor (circled in yellow). Scale bar: 3 µm. c Examples of TF-YPet interphase localization in NIH-3T3 cells, as compared to Hoechst staining and ranked by mitotic bound fraction (for n values see Supplementary Table 2). Bra Brachyury. Scale bar: 5 µm. d Correlation between the TF-YPet/Hoechst co-localization (MAFK: n = 8, NANOG and BHLHB8: n = 11, others: n = 10) and the mitotic bound fraction. e-g Correlation between the mitotic bound fraction and the fraction of YPet signal co-localized with heterochromatic regions (e), DNA-rich regions (f), and DNA-poor regions (g). n = 10. Error bars: SEM. r-values (r) and p-values (p) are based on Pearson correlation
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